N1-pyrazolospiroketone acetyl-CoA carboxylase inhibitors

ABSTRACT

The invention provides a compound of Formula (I) 
                         
or a pharmaceutically acceptable salt of the compound, wherein R 1 , R 2 , R 3  and R 4  are as described herein; pharmaceutical compositions thereof; and the use thereof in treating diseases, conditions or disorders modulated by the inhibition of an acetyl-CoA carboxylase enzyme(s) in an animal.

This application claims the benefit under 35 USC 119(e) to ProvisionalPatent Application No. 61/259,823 filed Nov. 10, 2009.

FIELD OF THE INVENTION

This invention relates to substituted pyrazolospiroketone compounds thatact as inhibitors of an acetyl-CoA carboxylase(s) and their use intreating diseases, conditions or disorders modulated by the inhibitionof acetyl-CoA carboxylase enzyme(s).

BACKGROUND OF THE INVENTION

Acetyl-CoA carboxylases (ACC) are a family of enzymes found in mostspecies and are associated with fatty acid synthesis and metabolismthrough catalyzing the production of malonyl-CoA from acetyl-CoA. Inmammals, two isoforms of the ACC enzyme have been identified. ACC1,which is expressed at high levels in lipogenic tissues, such as fat andthe liver, controls the first committed step in the biosynthesis oflong-chain fatty acids. If acetyl-CoA is not carboxylated to formmalonyl-CoA, it is metabolized through the Krebs cycle. ACC2, a minorcomponent of hepatic ACC but the predominant isoform in heart andskeletal muscle, catalyzes the production of malonyl-CoA at thecytosolic surface of mitochondria, and regulates how much fatty acid isutilized in β-oxidation by inhibiting carnitine palmitoyl transferase.Thus, by increasing fatty acid utilization and by preventing increasesin de novo fatty acid synthesis, chronic administration of an ACCinhibitor (ACC-I) may also deplete liver and adipose tissue triglyceride(TG) stores in obese subjects consuming a high or low-fat diet, leadingto selective loss of body fat.

Studies conducted by Abu-Etheiga, et al., suggest that ACC2 plays anessential role in controlling fatty acid oxidation and, as such it wouldprovide a target in therapy against obesity and obesity-relateddiseases, such as type-2 diabetes. See, Abu-Etheiga, L., et al.,“Acetyl-CoA carboxylase 2 mutant mice are protected against obesity anddiabetes induced by high-fat/high-carbohydrate diets” PNAS, 100(18)10207-10212 (2003). See also, Choi, C. S., et al., “Continuous fatoxidation in acetyl-CoA carboxylase 2 knockout mice increases totalenergy expenditure, reduces fat mass, and improves insulin sensitivity”PNAS, 104(42) 16480-16485 (2007).

It is becoming increasingly clear that hepatic lipid accumulation causeshepatic insulin resistance and contributes to the pathogenesis of type 2diabetes. Salvage, et al., demonstrated that ACC 1 and ACC2 are bothinvolved in regulating fat oxidation in hepatocytes while ACC1, thedominant isoform in rat liver, is the sole regulator of fatty acidsynthesis. Furthermore, in their model, combined reduction of bothisoforms is required to significantly lower hepatic malonyl-CoA levels,increase fat oxidation in the fed state, reduce lipid accumulation, andimprove insulin action in vivo. Thus, showing that hepatic ACC1 and ACC2inhibitors may be useful in the treatment of nonalcoholic fatty liverdisease (NAFLD) and hepatic insulin resistance. See, Savage, D. B., etal., “Reversal of diet-induced hepatic steatosis and hepatic insulinresistance by antisense oligonucleotide inhibitors of acetyl-CoAcarboxylases 1 and 2” J Clin Invest doi: 10.1172/JCI27300. See also, Oh,W., et al., “Glucose and fat metabolism in adipose tissue of acetyl-CoAcarboxylase 2 knockout mice” PNAS, 102(5) 1384-1389 (2005).

Consequently, there is a need for medicaments containing ACC1 and/orACC2 inhibitors to treat obesity and obesity-related diseases (such as,NAFLD and type-2 diabetes) by inhibiting fatty acid synthesis and byincreasing fatty acid oxidation.

SUMMARY OF THE INVENTION

The present invention relates to compounds having the structure ofFormula (I)

wherein R¹ is (C₁-C₆)alkyl, (C₃-C₇)cycloalkyl, tetrahydrofuranyl oroxetanyl; wherein said (C₁-C₆)alkyl is optionally substituted with 1 to2 substituents independently selected from (C₁-C₃)alkoxy; hydroxy, halo,phenyl, tetrahydrofuranyl or oxetanyl;

R² is hydrogen, halo, (C₁-C₃)alkyl, cyano or —C(═NH)(OCH₃);

R³ are each independently hydrogen or (C₁-C₃)alkyl;

R⁴ is (C₆-C₁₀)aryl, 5 to 12 membered heteroaryl or 8 to 12 memberedfused heterocyclicaryl; wherein said (C₆-C₁₀)aryl, 5 to 12 memberedheteroaryl or 8 to 12 membered fused heterocyclicaryl are eachoptionally substituted with one to three substituents independentlyselected from (C₁-C₃)alkyl, (C₁-C₃)alkoxy, halo, amino,(C₁-C₃)alkylamino, di(C₁-C₃)alkylamino, hydroxy, cyano, amido, phenyl, 5to 6 membered heteroaryl or 5 to 6 membered heterocyclyl; or apharmaceutically acceptable salt thereof.

A preferred embodiment of the present invention are compounds of Formula(I) wherein R⁴ is (C₆-C₁₀)aryl selected from phenyl or naphthyl; a 5 to12 membered heteroaryl selected from pyridinyl, pyrazolyl, pyrimidinyl,triazolyl, indolizinyl, indazolyl, pyrrolo[2,3-b]pyridinyl,pyrrolo[3,2-b]pyridinyl, pyrrolo[1,2-a]pyrazinyl,imidazo[1,2-a]pyridinyl, imidazo[1,5-a]pyridinyl, benzo[d]imidazolyl,pyrazolo[3,4-b]pyridinyl, pyrazolo[4,3-b]pyridinyl,pyrazolo[1,5-a]pyrimidinyl, benzo[d]imidazol-2-onyl, 1,6-naphthyridinyl,quinoxalinyl, quinolin-4-onyl or isoquinolin-1-onyl; or an 8 to 12membered fused heterocyclicaryl selected from 3,4-dihydroquinolin-2-onylor indolin-2-onyl; wherein each R⁴ group is optionally substituted withone to four substituents independently selected from (C₁-C₃)alkyl,(C₁-C₃)alkoxy, halo, amino, (C₁-C₃)alkylamino, di(C₁-C₃)alkylamino,hydroxy, cyano, amido, phenyl, 5 to 6 membered heteroaryl or 5 to 6membered heterocyclyl; or a pharmaceutically acceptable salt thereof.

Another preferred embodiment of the present invention is the compound ofFormula (I) wherein R¹ is (C₁-C₆)alkyl, (C₃-C₇)cycloalkyl, ortetrahydrofuranyl; and R² is hydrogen or methyl; or a pharmaceuticallyacceptable salt thereof. Yet another preferred embodiment of the presentinvention is the compound of formula (I) wherein R¹ is ethyl, isopropylor t-butyl; and R⁴ is phenyl, pyrazolyl, imidazolyl, triazolyl,pyridinyl, pyrimidinyl, indolyl, benzopyrazinyl, benzoimidazolyl,benzoimidazolonyl, pyrrolopyridinyl, pyrrolopyrimidinyl,pyrazolopyridinyl, pyrazolopyrimidinyl, indazolyl, indolinonyl,naphthyridinyl, quinolinyl, quinolinonyl, dihydroquinolinonyl,oxo-dihydroquinolinonyl, isoquinolinyl, isoquinolinonyl,dihydroisoquinonyl or oxo-dihydroisoquinonyl, each optionallysubstituted with one to three substituents independently selected fromfluoro, chloro, methyl, amino, methylamino, dimethylamino, amido, cyano,phenyl, imidazolyl, pyrazolyl, triazolyl, pyridinyl or morpholinyl; or apharmaceutically acceptable salt thereof. A further preferred embodimentof the present invention is the compound of formula (I) wherein R¹ isisopropyl or t-butyl; R² is hydrogen; and each R³ is hydrogen; or apharmaceutically acceptable salt thereof. Yet another preferredembodiment of the present invention is the compound of formula (I)wherein R⁴ is indazolyl, benzoimidazolyl,1-oxo-1,2-dihydroisoquinolinyl, 1H-pyrrolo[3,2-b]pyridinyl,2-oxo-2,3-dihydro-1H-benzo[d]imidazolyl, 1H-pyrazolylphenyl,1H-pyrazolylpyridinyl, or 1H-imidazolylphenyl; each optionallysubstituted with one to two methyl, chloro or fluoro; or apharmaceutically acceptable salt thereof.

Another preferred embodiment of the present invention is a compoundselected from1-isopropyl-1′-(1-oxo-1,2-dihydroisoquinoline-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(1H-pyrrolo[3,2-b]pyridine-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(2-methyl-3H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(1H-pyrrolo[3,2-b]pyridine-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(1H-indazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(7-fluoro-1H-indazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(1-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(7-chloro-2-methyl-3H-benzo[d]imidazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(1H-indazole-6-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(3-(1H-pyrazol-4-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(2-methyl-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H-one;1′-(1H-indazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(1H-indazole-5-carbonyl)-1-isopropyl-3-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(2-(1H-pyrazol-3-yl)pyridine-4-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(3-(1H-pyrazol-3-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(1H-pyrrolo[3,2-b]pyridine-2-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(4-(1H-imidazol-2-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(3-(1H-imidazol-2-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;or1-(tert-butyl)-1′-(1H-indazole-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;or a pharmaceutically acceptable salt thereof.

Another aspect of the present invention is a pharmaceutical compositioncomprising an amount of a compound of formula (I) as described in any ofthe embodiments; or a pharmaceutically acceptable salt thereof and apharmaceutically acceptable excipient, diluent, or carrier. Preferably,the composition comprises a therapeutically effective amount of acompound of the present invention or a pharmaceutically acceptable saltthereof. The composition may also contain at least one additionalpharmaceutical agent. Preferred agents include anti-diabetic agentsand/or anti-obesity agents (described herein below).

In yet another aspect of the present invention is a method for treatinga disease, condition, or disorder mediated by the inhibition ofacetyl-CoA carboxylase enzyme(s) in a mammal that includes the step ofadministering to a mammal, preferably a human, in need of such treatmenta therapeutically effective amount of a compound of the presentinvention or a pharmaceutically acceptable salt thereof, or apharmaceutical composition thereof.

Diseases, disorders, or conditions mediated by inhibitors of acetyl-CoAcarboxylases include Type II diabetes and diabetes-related diseases,such as nonalcoholic fatty liver disease (NAFLD), hepatic insulinresistance, hyperglycemia, metabolic syndrome, impaired glucosetolerance, diabetic neuropathy, diabetic nephropathy, diabeticretinopathy, obesity, dyslipidemia, hypertension, hyperinsulinemia, andinsulin resistance syndrome. Preferred diseases, disorders, orconditions include Type II diabetes, nonalcoholic fatty liver disease(NAFLD), hepatic insulin resistance, hyperglycemia, impaired glucosetolerance, obesity, and insulin resistance syndrome. More preferred areType II diabetes, nonalcoholic fatty liver disease (NAFLD), hepaticinsulin resistance, hyperglycemia, and obesity. Most preferred is TypeII diabetes.

A preferred embodiment is a method for treating, (e.g. delaying theprogression or onset) of Type 2 diabetes and diabetes-related disordersin animals comprising the step of administering to an animal in need ofsuch treatment a therapeutically effective amount of a compound of thepresent invention or a pharmaceutically acceptable salt thereof or acomposition thereof.

Another preferred embodiment is a method for treating obesity andobesity-related disorders in animals comprising the step ofadministering to an animal in need of such treatment a therapeuticallyeffective amount of a compound of the present invention or apharmaceutically acceptable salt thereof or a composition thereof.

Yet another preferred embodiment is a method for treating nonalcoholicfatty liver disease (NAFLD) or hepatic insulin resistance in animalscomprising the step of administering to an animal in need of suchtreatment a therapeutically effective amount of a compound of thepresent invention or a pharmaceutically acceptable salt thereof or acomposition thereof.

Compounds of the present invention may be administered in combinationwith other pharmaceutical agents (in particular, anti-obesity andanti-diabetic agents described herein below). The combination therapymay be administered as (a) a single pharmaceutical composition whichcomprises a compound of the present invention or a pharmaceuticallyacceptable salt thereof, at least one additional pharmaceutical agentdescribed herein and a pharmaceutically acceptable excipient, diluent,or carrier; or (b) two separate pharmaceutical compositions comprising(i) a first composition comprising a compound of the present inventionor a pharmaceutically acceptable salt thereof and a pharmaceuticallyacceptable excipient, diluent, or carrier, and (ii) a second compositioncomprising at least one additional pharmaceutical agent described hereinand a pharmaceutically acceptable excipient, diluent, or carrier. Thepharmaceutical compositions may be administered simultaneously orsequentially and in any order.

SEQ. ID NO. 1 provides a sequence of recombinant human ACC1 (SEQ. IDNO. 1) that can be employed in the Transcreener in vitro assay.

SEQ. ID NO. 2 provides a sequence of recombinant human ACC2 (SEQ. ID NO.2) that can be employed in the Transcreener in vitro assay.

DETAILED DESCRIPTION OF THE INVENTION Definitions

The phrase “therapeutically effective amount” means an amount of acompound of the present invention or a pharmaceutically acceptable saltthereof that: (i) treats or prevents the particular disease, condition,or disorder, (ii) attenuates, ameliorates, or eliminates one or moresymptoms of the particular disease, condition, or disorder, or (iii)prevents or delays the onset of one or more symptoms of the particulardisease, condition, or disorder described herein.

The term “animal” refers to humans (male or female), companion animals(e.g., dogs, cats and horses), food-source animals, zoo animals, marineanimals, birds and other similar animal species. “Edible animals” refersto food-source animals such as cows, pigs, sheep and poultry.

The phrase “pharmaceutically acceptable” indicates that the substance orcomposition must be compatible chemically and/or toxicologically, withthe other ingredients comprising a formulation, and/or the mammal beingtreated therewith.

The terms “treating”, “treat”, or “treatment” embrace both preventative,i.e., prophylactic, and palliative treatment.

The terms “modulated” or “modulating”, or “modulate(s)”, as used herein,unless otherwise indicated, refers to the inhibition of the Acetyl-CoAcarboxylases (ACC) enzyme(s) with compounds of the present invention.

The terms “mediated” or “mediating” or “mediate(s)”, as used herein,unless otherwise indicated, refers to the (i) treatment or preventionthe particular disease, condition, or disorder, (ii) attenuation,amelioration, or elimination of one or more symptoms of the particulardisease, condition, or disorder, or (iii) prevention or delay of theonset of one or more symptoms of the particular disease, condition, ordisorder described herein, by inhibiting the Acetyl-CoA carboxylases(ACC) enzyme(s).

The term “compounds of the present invention” (unless specificallyidentified otherwise) refer to compounds of Formula (I) and anypharmaceutically acceptable salts of the compounds, as well as, allstereoisomers (including diastereoisomers and enantiomers), tautomers,conformational isomers, and isotopically labeled compounds. Hydrates andsolvates of the compounds of the present invention are consideredcompositions of the present invention, wherein the compound is inassociation with water or solvent, respectively.

The terms “(C₁-C₆)alkyl” and “(C₁-C₃)alkyl” are alkyl groups of thespecified number of carbons, from one to six or one to three carbons,respectively, which can be either straight chain or branched. Forexample, the term “(C₁-C₃)alkyl” has from one to three carbons andconsists of methyl, ethyl, n-propyl and isopropyl.

The term “(C₃-C₇)cycloalkyl” means a cycloalkyl group with three toseven carbon atoms and consists of cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl and cycloheptyl. The term “halo” means fluoro, chloro, bromoor iodo. The term “(C₆-C₁₀)aryl” means an aromatic carbocyclic groupconsisting of six to ten carbon atoms such as phenyl or naphthyl.

The term “5 to 12 membered heteroaryl” means a five to twelve memberedaromatic group which contains at least one heteroatom selected fromnitrogen, oxygen and sulfur. As used herein the point of attachment ofthe “5 to 12 membered heteroaryl” group is on a carbon atom of thatgroup. The “5 to 12 membered heteroaryl” group can be either monocyclicor bicyclic. Preferred embodiments of monocyclic heteroaryls include,but are not limited to, pyrazolyl, imidazolyl, triazolyl, pyridinyl, andpyrimidinyl. Preferred embodiments of bicyclic heteroaryls include, butare not limited to, radicals of the following ring systems:

The term “8 to 12 membered fused heterocyclicaryl” means an 8 to 12membered ring system in which a non-aromatic heterocyclic ring is fusedto an aryl ring. As used herein the point of attachment of the “8 to 12membered fused heterocyclicaryl” group is on a carbon atom of thatgroup. A preferred embodiment includes radicals of ring systems such as:

Compounds of the present invention may be synthesized by syntheticroutes that include processes analogous to those well-known in thechemical arts, particularly in light of the description containedherein. The starting materials are generally available from commercialsources such as Aldrich Chemicals (Milwaukee, Wis.) or are readilyprepared using methods well known to those skilled in the art (e.g.,prepared by methods generally described in Louis F. Fieser and MaryFieser, Reagents for Organic Synthesis, v. 1-19, Wiley, New York(1967-1999 ed.), or Beilsteins Handbuch der organischen Chemie, 4, Aufl.ed. Springer-Verlag, Berlin, including supplements (also available viathe Beilstein online database)).

For illustrative purposes, the reaction schemes depicted below providepotential routes for synthesizing the compounds of the present inventionas well as key intermediates. For a more detailed description of theindividual reaction steps, see the Examples section below. Those skilledin the art will appreciate that other synthetic routes may be used tosynthesize the inventive compounds. Although specific starting materialsand reagents are depicted in the schemes and discussed below, otherstarting materials and reagents can be easily substituted to provide avariety of derivatives and/or reaction conditions. In addition, many ofthe compounds prepared by the methods described below can be furthermodified in light of this disclosure using conventional chemistry wellknown to those skilled in the art.

In the preparation of compounds of the present invention, protection ofremote functionality (e.g., primary or secondary amine) of intermediatesmay be necessary. The need for such protection will vary depending onthe nature of the remote functionality and the conditions of thepreparation methods. Suitable amino-protecting groups (NH-Pg) includeacetyl, trifluoroacetyl, t-butoxycarbonyl (BOC), benzyloxycarbonyl (CBz)and 9-fluorenylmethyleneoxycarbonyl (Fmoc). Similarly, a“hydroxy-protecting group” refers to a substituent of a hydroxy groupthat blocks or protects the hydroxy functionality. Suitablehydroxyl-protecting groups (O-Pg) include for example, allyl, acetyl,silyl, benzyl, para-methoxybenzyl, trityl, and the like. The need forsuch protection is readily determined by one skilled in the art. For ageneral description of protecting groups and their use, see T. W.Greene, Protective Groups in Organic Synthesis, John Wiley & Sons, NewYork, 1991.

The following reaction schemes, Reaction Scheme I through ReactionScheme IV provide representative procedures that are used to prepare thecompounds of Formula (I). It is to be understood that these reactionschemes are to be construed in a non-limiting manner and that reasonablevariations of the depicted methods can be used to prepare the compoundsof Formula (I).

Reaction Scheme I outlines the general procedures one could use toprovide compounds of the present invention having Formula (Ia) which arecompounds of Formula (I) in which R² and each R³ are each hydrogen. Theprotected spiropiperidine derivative (VIIIa) may be formed by treatingthe appropriately protected piperidine aldehyde (Xa) with methyl vinylketone (IXa). The group Pg represents an appropriate amine protectinggroup and is preferably N-tert-butoxycarbonyl (BOG) or carbobenzyloxy(Cbz). This reaction can be carried out in the presence of ethanolicpotassium hydroxide according to a procedure analogous to that describedby Roy, S. et al., Chem. Eur. J. 2006, 12, 3777-3788 at 3786.Alternatively, the reaction can be carried out in the presence ofpara-toluenesulfonic acid (pTSA) in refluxing benzene to provide thedesired product (VIIIa).

The spiropiperidine derivative (VIIIa) can then be reacted withtris-(N,N-dimethylamino)methane in refluxing toluene to provide theenamine functionalized spiropiperidine derivative (VIIa). Alternatively,the enamine (VIIa) may be prepared by reacting spiropiperidine (VIIIa)with N,N-dimethylformamide dimethyl acetal as solvent at reflux. Thisreaction may also be run in an alcoholic solvent such as 2-propanol, anaromatic hydrocarbon solvent such as toluene or a polar aprotic solventsuch as N,N-dimethylformamide. Additionally, this reaction may becatalyzed by the addition of 4-toluenesulfonic acid,tris(dimethylamino)methane, or various bases such as lithium hydroxide,DBU and N,N-diisopropylethylamine. This transformation may also beperformed with, or by reaction with, t-butoxy bis(dimethylamino)methanein toluene at reflux.

Compound (VIIa) is then reacted with an appropriate hydrazine derivativeR¹NHNH² in the presence of acetic acid in refluxing ethanol or tolueneto provide the desired cyclized compound of formula (VIa) (see MuraliDhar, T. G. et al. Bioorg. Med. Chem. Lett. 2007, 17, 5019-5024 at5020). The compound of formula (VIa) can then be treated withN-bromosuccinimide (NBS) in the presence of water in THF to provide thecorresponding bromo hydroxy derivative of formula (Va). The bromohydroxy derivative (Va) is then oxidized with Jones reagent in a methodanalogous to that provided in Wolinsky, J. et al., J. Org. Chem. 1978,43(5), 875-881 at 876, 879 to provide the α-bromo keto derivative offormula (IVa). Alternatively the oxidation of (Va) can be performed withcatalytic tetrapropylammonium perruthenate and N-methylmorpholineN-oxide. The compound of formula (IVa) can then be debrominated usingconventional methods such as treatment with zinc and acetic acid or,alternatively, zinc in the presence of aqueous ammonium chloride intetrahydrofuran to provide the compound of formula (IIIa).

The compound of formula (IIIa) can then be deprotected to provide thefree spiropiperidine derivative of formula (IIa) using standard methodswhich depend on which protecting group Pg has been employed. Forexample, when Pg represents tert-butyloxycarbonyl (BOC) standard strongacid deprotection conditions such as 4N hydrochloric acid in dioxane ortrifluoroacetic acid in an appropriate solvent such as dichloromethanecan be used to remove the BOC group. When Pg represents carbobenzyloxy(Cbz), hydrogenation over palladium on carbon in ethanol or treatmentwith a hydrogen source such as ammonium formate or1-methyl-1,4-cyclohexadiene in the presence of palladium on carbon inethanol or ethyl acetate can be employed to carry out the deprotection.

The spiropiperidine derivative of Formula (IIa) can then be acylated byemploying standard methods to provide the compound of Formula (Ia). Forexample, the compound (Ia) may then be formed using a standard peptidecoupling reaction with the desired carboxylic acid (R⁴CO₂H). Forexample, the spiropiperidine intermediate (IIa) and carboxylic acid(R⁴CO₂H) may be coupled by forming an activated carboxylic acid ester,such as by contacting the carboxylic acid (R⁴CO₂H) with a peptidecoupling reagent, such asO-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (HATU) or1-ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC.HCl),in the presence or absence of an activating agent, such ashydroxybenzotriazole (HOBt) and in the presence of a suitable base, suchas N,N-diisopropylethylamine (DIEA), triethylamine or N-methylmorpholine(NMM), in a suitable solvent such as THF and/or DMF, dimethylacetamide(DMA) or dichloromethane and then contacting the activated carboxylicacid ester with the spiropiperidine derivative (IIa) to form a compoundof Formula (Ia).

Alternatively, compounds of Formula (Ia) can be formed by firstconverting the carboxylic acid (R⁴CO₂H) to an acid chloride (R⁴COCl),such as by reacting with thionyl chloride, and then reacting the acidchloride with the spiropiperidine derivative (IIa) in the presence of anappropriate base such as triethylamine in an appropriate solvent such asdichloromethane to form a compound of Formula (Ia). Still anotheralternative method entails treating the carboxylic acid (R⁴CO₂H) with2-chloro-4,6-dimethoxytriazine in the presence of a suitable base, suchas N-methylmorpholine in a suitable solvent such as THF and/or DMF. Tothe activated ester is added a solution of the spiropiperidinederivative (IIa) and base, such as N-methylmorpholine, in a suitablesolvent, such as THF and/or DMF which then provides the compound ofFormula (Ia).

Reaction Scheme II provides an alternative synthesis of compounds ofFormula (Ia) starting from the intermediate of Formula (VIa). Thecompound of Formula (VIa) is treated with N-bromosuccinimide (NBS) inthe presence of methanol in THF, or preferably in methanol, (Nishimura,T. et al. Org. Lett. 2008, 10(18), 4057-4060 at 4059) to provide themethoxy bromo spiropiperidine derivative of Formula (Vb). Base inducedelimination of the compound of Formula (Vb) by treatment with a strongbase such as potassium tert-butoxide in THF provides the compound ofFormula (IVb) which is then treated with a strong acid such as 2Nhydrochloric acid in THF to provide the compound of Formula (IIIa). Thecompound of Formula (IIIa) can then be deprotected and acylated asdescribed previously in Reaction Scheme I to provide compounds ofFormula (Ia).

Reaction Scheme III provides a synthesis of compounds of Formula (Ib)which are compounds of Formula (I) in which R² is bromo and each R³ ishydrogen. The compound of Formula (VIa) is reacted with approximatelytwo equivalents of N-bromosuccinimide in the presence of methanol toprovide the dibromo methoxy spiropiperidine derivative of Formula (Vc).The compound of Formula (Vc) is then subjected to elimination conditionsby treatment with a strong base such as potassium tert-butoxide in anappropriate solvent to provide the compound of Formula (IVc). Treatmentof the compound of formula (IVc) with strong acid such as 2Nhydrochloric acid provides the compound of Formula (IIIb). Deprotectionof the compound of Formula (IIIb) to provide the compound of Formula(IIb) followed by acylation to provide the compound of Formula (Ib) canbe carried out as described previously for Reaction Scheme I.

Reaction Scheme IV depicts the preparation of certain other compoundswithin Formula (I) from certain of the intermediates previouslydepicted. The first transformation in Reaction Scheme IV showsintroduction of a methyl group at the R² position by reacting the bromospiropiperidine derivative of Formula (IIIb) with trimethylboroxine indimethylformamide in the presence of an appropriate palladium catalyst,such as palladium tetrakis triphenylphosphine in the presence ofpotassium carbonate and water to provide the compound of Formula (IIIc).Other alkyl groups can be introduced at the R² position in an analogousmanner. The compound of Formula (IIIc) can then be deprotected andacylated as previously described. The second transformation in ReactionScheme IV depicts the introduction of a cyano group at the R² position.The bromo spiropiperidine compound (IIIb) is reacted with zinc cyanidein the presence of zinc and an appropriate palladium catalyst to providethe compound of Formula (IIId) which can then be deprotected andacylated to provide a compound of Formula (Id). The third transformationin Reaction Scheme IV depicts introduction of an appropriate group atthe R³ position of the compound of Formula (IIIe). The compound ofFormula (IIIe) is deprotonated with a strong base, such as lithiumhexamethyldisilazide (LHMDS) under appropriate anhydrous conditions inan appropriate solvent, preferably at low temperature. The enolate thusformed is then reacted with an appropriate electrophile R³Lg wherein Lgrepresents an appropriate leaving group (such as a halide when R³Lg isan alkyl halide) to provide the compound of Formula (IIIf) wherein R³ isan appropriate group such as an alkyl group. The deprotonation of thecompound of Formula (IIIf) and reaction with another R³Lg can then becarried out again if desired. The compound of Formula (IIIf) can then bedeprotected and acylated as previously described to provide the compoundof Formula (Ie).

The compounds of the present invention may be isolated and used per seor in the form of their pharmaceutically acceptable salts. In accordancewith the present invention, compounds with multiple basic nitrogen atomscan form salts with varying number of equivalents (“eq.”) of acid. Itwill be understood by practitioners that all such salts are within thescope of the present invention.

Pharmaceutically acceptable salts, as used herein in relation tocompounds of the present invention, include pharmaceutically acceptableinorganic and organic salts of the compound. These salts can be preparedin situ during the final isolation and purification of a compound, or byseparately reacting the compound thereof, with a suitable organic orinorganic acid and isolating the salt thus formed. Representative saltsinclude, but are not limited to, the hydrobromide, hydrochloride,hydroiodide, sulfate, bisulfate, nitrate, acetate, trifluoroacetate,oxalate, besylate, palmitate, pamoate, malonate, stearate, laurate,malate, borate, benzoate, lactate, phosphate, hexafluorophosphate,benzene sulfonate, tosylate, formate, citrate, maleate, fumarate,succinate, tartrate, naphthylate, mesylate, glucoheptonate, lactobionateand laurylsulphonate salts, and the like. These may also include cationsbased on the alkali and alkaline earth metals, such as sodium, lithium,potassium, calcium, magnesium, and the like, as well as non-toxicammonium, quaternary ammonium, and amine cations including, but notlimited to, ammonium, tetramethylammonium, tetraethylammonium,methylammonium, dimethylammonium, trimethylammonium, triethylammonium,ethylammonium, and the like. For additional examples see, for example,Berge, et al., J. Pharm. Sci., 66, 1-19 (1977).

Compounds of the present invention may exist in more than one crystalform. Polymorphs of compounds of Formula (I) and salts thereof(including solvates and hydrates) form part of this invention and may beprepared by crystallization of a compound of the present invention underdifferent conditions. For example, using different solvents or differentsolvent mixtures for recrystallization; crystallization at differenttemperatures; various modes of cooling, ranging from very fast to veryslow cooling during crystallization. Polymorphs may also be obtained byheating or melting a compound of the present invention followed bygradual or fast cooling. The presence of polymorphs may be determined bysolid probe nuclear magnetic resonance (NMR) spectroscopy, infrared (IR)spectroscopy, differential scanning calorimetry, powder X-raydiffraction or such other techniques.

This invention also includes isotopically-labeled compounds, which areidentical to those described by Formula (1), but for the fact that oneor more atoms are replaced by an atom having an atomic mass or massnumber different from the atomic mass or mass number usually found innature. Examples of isotopes that can be incorporated into compounds ofthe invention include isotopes of hydrogen, carbon, nitrogen, oxygen,sulfur and fluorine, such as ²H, ³H, ¹³C, ¹⁵N, ¹⁸O, ¹⁷O, ³⁵S, ³⁶Cl,¹²⁵I, ¹²⁹I, and ¹⁸F respectively. Certain isotopically-labeled compoundsof the present invention, for example those into which radioactiveisotopes such as ³H and ¹⁴C are incorporated, are useful in drug and/orsubstrate tissue distribution assays. Tritiated (i.e., ³H), andcarbon-14 (i.e., ¹⁴C), isotopes are particularly preferred for theirease of preparation and detectability. Further, substitution withheavier isotopes such as deuterium (i.e., ²H), can afford certaintherapeutic advantages resulting from greater metabolic stability, forexample increased in vivo half-life or reduced dosage requirements and,hence, may be preferred in some circumstances. Isotopically labeledcompounds of the present invention can generally be prepared by carryingout the procedures disclosed in the schemes and/or in the Examplesbelow, by substituting a readily available isotopically labeled reagentfor a non-isotopically labeled reagent.

The compounds of the present invention may contain stereogenic centers.These compounds may exist as mixtures of enantiomers or as pureenantiomers. When the compound includes a stereogenic center, thecompound may be resolved into the pure enantiomers by methods known tothose skilled in the art, for example by formation of diastereoisomericsalts which may be separated, for example, by crystallization; formationof stereoisomeric derivatives or complexes which may be separated, forexample, by crystallization, gas-liquid or liquid chromatography;selective reaction of one enantiomer with an enantiomer-specificreagent, for example enzymatic esterification; or gas-liquid or liquidchromatography in a chiral environment, for example on a chiral supportfor example silica with a bound chiral ligand or in the presence of achiral solvent. It will be appreciated that where the desiredstereoisomer is converted into another chemical entity by one of theseparation procedures described above, a further step is required toliberate the desired enantiomeric form. Alternatively, the specificstereoisomers may be synthesized by using an optically active startingmaterial, by asymmetric synthesis using optically active reagents,substrates, catalysts or solvents, or by converting one stereoisomerinto the other by asymmetric transformation.

Compounds of the present invention may exist in different stableconformational forms which may be separable. Torsional asymmetry due torestricted rotation about an asymmetric single bond, for example becauseof steric hindrance or ring strain, may permit separation of differentconformers. The compounds of the present invention further include eachconformational isomer of compounds of Formula (I) and mixtures thereof.

Compounds of the present invention are useful for treating diseases,conditions and/or disorders modulated by the inhibition of theacetyl-CoA carboxylases enzyme(s) (in particular, ACC1 and ACC2).Another embodiment of the present invention is a pharmaceuticalcomposition comprising a therapeutically effective amount of a compoundof the present invention or a pharmaceutically acceptable salt thereofand a pharmaceutically acceptable excipient, diluent or carrier. Thecompounds of the present invention (including the compositions andprocesses used therein) may also be used in the manufacture of amedicament for the therapeutic applications described herein.

A typical formulation is prepared by mixing a compound of the presentinvention and a carrier, diluent or excipient. Suitable carriers,diluents and excipients are well known to those skilled in the art andinclude materials such as carbohydrates, waxes, water soluble and/orswellable polymers, hydrophilic or hydrophobic materials, gelatin, oils,solvents, water, and the like. The particular carrier, diluent orexcipient used will depend upon the means and purpose for which thecompound of the present invention is being applied. Solvents aregenerally selected based on solvents recognized by persons skilled inthe art as safe (GRAS) to be administered to a mammal. In general, safesolvents are non-toxic aqueous solvents such as water and othernon-toxic solvents that are soluble or miscible in water. Suitableaqueous solvents include water, ethanol, propylene glycol, polyethyleneglycols (e.g., PEG400, PEG300), etc. and mixtures thereof. Theformulations may also include one or more buffers, stabilizing agents,surfactants, wetting agents, lubricating agents, emulsifiers, suspendingagents, preservatives, antioxidants, opaquing agents, glidants,processing aids, colorants, sweeteners, perfuming agents, flavoringagents and other known additives to provide an elegant presentation ofthe drug (i.e., a compound of the present invention or apharmaceutically acceptable salt thereof or a pharmaceutical compositionthereof) or aid in the manufacturing of the pharmaceutical product(i.e., for use in medicine such as preparing a medicament).

The formulations may be prepared using conventional dissolution andmixing procedures. For example, the bulk drug substance (i.e., compoundof the present invention or stabilized form of the compound (e.g.,complex with a cyclodextrin derivative or other known complexationagent)) is dissolved in a suitable solvent in the presence of one ormore of the excipients described above. The dissolution rate of poorlywater-soluble compounds may be enhanced by the use of a spray-drieddispersion, such as those described by Takeuchi, H., et al. in“Enhancement of the dissolution rate of a poorly water-soluble drug(tolbutamide) by a spray-drying solvent deposition method anddisintegrants” J. Pharm. Pharmacol., 39, 769-773 (1987); and EP0901786B1 (US2002/009494), incorporated herein by reference. The compound ofthe present invention is typically formulated into pharmaceutical dosageforms to provide an easily controllable dosage of the drug and to givethe patient an elegant and easily handleable product.

The pharmaceutical compositions also include solvates and hydrates ofthe compounds of the present invention. The term “solvate” refers to amolecular complex of a compound represented by Formula (I) (includingpharmaceutically acceptable salts thereof) with one or more solventmolecules. Such solvent molecules are those commonly used in thepharmaceutical art, which are known to be innocuous to the recipient,e.g., water, ethanol, ethylene glycol, and the like, The term “hydrate”refers to the complex where the solvent molecule is water. The solvatesand/or hydrates preferably exist in crystalline form. Other solvents maybe used as intermediate solvates in the preparation of more desirablesolvates, such as methanol, methyl t-butyl ether, ethyl acetate, methylacetate, (S)-propylene glycol, (R)-propylene glycol, 1,4-butyne-diol,and the like.

The pharmaceutical composition (or formulation) for application may bepackaged in a variety of ways depending upon the method used foradministering the drug. Generally, an article for distribution includesa container having deposited therein the pharmaceutical formulation inan appropriate form. Suitable containers are well-known to those skilledin the art and include materials such as bottles (plastic and glass),sachets, ampoules, plastic bags, metal cylinders, and the like. Thecontainer may also include a tamper-proof assemblage to preventindiscreet access to the contents of the package. In addition, thecontainer has deposited thereon a label that describes the contents ofthe container. The label may also include appropriate warnings.

The present invention further provides a method of treating diseases,conditions and/or disorders modulated by the inhibition of theacetyl-CoA carboxylases enzyme(s) in an animal that includesadministering to an animal in need of such treatment a therapeuticallyeffective amount of a compound of the present invention or apharmaceutical composition comprising an effective amount of a compoundof the present invention and a pharmaceutically acceptable excipient,diluent, or carrier. The method is particularly useful for treatingdiseases, conditions and/or disorders that benefit from the inhibitionof acetyl-CoA carboxylases enzyme(s).

One aspect of the present invention is the treatment of obesity, andobesity-related disorders (e.g., overweight, weight gain, or weightmaintenance). Obesity and overweight are generally defined by body massindex (BMI), which is correlated with total body fat and estimates therelative risk of disease. BMI is calculated by weight in kilogramsdivided by height in meters squared (kg/m²). Overweight is typicallydefined as a BMI of 25-29.9 kg/m², and obesity is typically defined as aBMI of 30 kg/m². See, e.g., National Heart, Lung, and Blood Institute,Clinical Guidelines on the Identification, Evaluation, and Treatment ofOverweight and Obesity in Adults, The Evidence Report, Washington, D.C.:U.S. Department of Health and Human Services, NIH publication no.98-4083 (1998).

Another aspect of the present invention is for the treatment or delayingthe progression or onset of diabetes or diabetes-related disordersincluding Type 1 (insulin-dependent diabetes mellitus, also referred toas “IDDM”) and Type 2 (noninsulin-dependent diabetes mellitus, alsoreferred to as “NIDDM”) diabetes, impaired glucose tolerance, insulinresistance, hyperglycemia, and diabetic complications (such asatherosclerosis, coronary heart disease, stroke, peripheral vasculardisease, nephropathy, hypertension, neuropathy, and retinopathy).

In yet another aspect of the present invention is the treatment ofobesity co-morbidities, such as metabolic syndrome. Metabolic syndromeincludes diseases, conditions or disorders such as dyslipidemia,hypertension, insulin resistance, diabetes (e.g., Type 2 diabetes),coronary artery disease and heart failure. For more detailed informationon Metabolic Syndrome, see, e.g., Zimmet, P. Z., et al., “The MetabolicSyndrome: Perhaps an Etiologic Mystery but Far From a Myth—Where Doesthe International Diabetes Federation Stand?,” Diabetes & Endocrinology,7(2), (2005); and Alberti, K. G., et al., “The Metabolic Syndrome—A NewWorldwide Definition,” Lancet, 366, 1059-62 (2005). Preferably,administration of the compounds of the present invention provides astatistically significant (p<0.05) reduction in at least onecardiovascular disease risk factor, such as lowering of plasma leptin,C-reactive protein (CRP) and/or cholesterol, as compared to a vehiclecontrol containing no drug. The administration of compounds of thepresent invention may also provide a statistically significant (p<0.05)reduction in glucose serum levels.

In yet another aspect of the invention is the treatment of nonalcoholicfatty liver disease (NAFLD) and hepatic insulin resistance.

For a normal adult human having a body weight of about 100 kg, a dosagein the range of from about 0.001 mg to about 10 mg per kilogram bodyweight is typically sufficient, preferably from about 0.01 mg/kg toabout 5.0 mg/kg, more preferably from about 0.01 mg/kg to about 1 mg/kg.However, some variability in the general dosage range may be requireddepending upon the age and weight of the subject being treated, theintended route of administration, the particular compound beingadministered and the like. The determination of dosage ranges andoptimal dosages for a particular patient is well within the ability ofone of ordinary skill in the art having the benefit of the instantdisclosure. It is also noted that the compounds of the present inventioncan be used in sustained release, controlled release, and delayedrelease formulations, which forms are also well known to one of ordinaryskill in the art.

The compounds of the present invention may also be used in conjunctionwith other pharmaceutical agent(s) for the treatment of the diseases,conditions and/or disorders described herein. Therefore, methods oftreatment that include administering compounds of the present inventionin combination with other pharmaceutical agents are also provided.Suitable pharmaceutical agents that may be used in combination with thecompounds of the present invention include anti-obesity agents(including appetite suppressants), anti-diabetic agents,anti-hyperglycemic agents, lipid lowering agents, and anti-hypertensiveagents.

Suitable anti-obesity agents include 11β-hydroxy steroid dehydrogenase-1(11β-HSD type 1) inhibitors, stearoyl-CoA desaturase-1 (SCD-1)inhibitor, MCR-4 agonists, cholecystokinin-A (CCK-A) agonists, monoaminereuptake inhibitors (such as sibutramine), sympathomimetic agents, β₃adrenergic agonists, dopamine agonists (such as bromocriptine),melanocyte-stimulating hormone analogs, 5HT2c agonists, melaninconcentrating hormone antagonists, leptin (the OB protein), leptinanalogs, leptin agonists, galanin antagonists, lipase inhibitors (suchas tetrahydrolipstatin, i.e. orlistat), anorectic agents (such as abombesin agonist), neuropeptide-Y antagonists (e.g., NPY Y5antagonists), PYY₃₋₃₆ (including analogs thereof), thyromimetic agents,dehydroepiandrosterone or an analog thereof, glucocorticoid agonists orantagonists, orexin antagonists, glucagon-like peptide-1 agonists,ciliary neurotrophic factors (such as Axokine™ available from RegeneronPharmaceuticals, Inc., Tarrytown, N.Y. and Procter & Gamble Company,Cincinnati, Ohio), human agouti-related protein (AGRP) inhibitors,ghrelin antagonists, histamine 3 antagonists or inverse agonists,neuromedin U agonists, MTP/ApoB inhibitors (e.g., gut-selective MTPinhibitors, such as dirlotapide), opioid antagonist, orexin antagonist,and the like.

Preferred anti-obesity agents for use in the combination aspects of thepresent invention include gut-selective MTP inhibitors (e.g.,dirlotapide, mitratapide and implitapide, R56918 (CAS No. 403987) andCAS No. 913541-47-6), CCKa agonists (e.g.,N-benzyl-2-[4-(1H-indol-3-ylmethyl)-5-oxo-1-phenyl-4,5-dihydro-2,3,6,10b-tetraaza-benzo[e]azulen-6-yl]-N-isopropyl-acetamidedescribed in PCT Publication No. WO 2005/116034 or US Publication No.2005-0267100 A1), 5HT2c agonists (e.g., lorcaserin), MCR4 agonist (e.g.,compounds described in U.S. Pat. No. 6,818,658), lipase inhibitor (e.g.,Cetilistat), PYY₃₋₃₆ (as used herein “PYY₃₋₃₆” includes analogs, such aspeglated PYY₃₋₃₆ e.g., those described in US Publication 2006/0178501),opioid antagonists (e.g., naltrexone), oleoyl-estrone (CAS No.180003-17-2), obinepitide (TM30338), pramlintide (Symlin®), tesofensine(NS2330), leptin, liraglutide, bromocriptine, orlistat, exenatide(Byetta®), AOD-9604 (CAS No. 221231-10-3) and sibutramine. Preferably,compounds of the present invention and combination therapies areadministered in conjunction with exercise and a sensible diet.

Suitable anti-diabetic agents include a sodium-glucose co-transporter(SGLT) inhibitor, a phosphodiesterase (PDE)-10 inhibitor, adiacylglycerol acyltransferase (DGAT) 1 or 2 inhibitor, a sulfonylurea(e.g., acetohexamide, chlorpropamide, diabinese, glibenclamide,glipizide, glyburide, glimepiride, gliclazide, glipentide, gliquidone,glisolamide, tolazamide, and tolbutamide), a meglitinide, an α-amylaseinhibitor (e.g., tendamistat, trestatin and AL-3688), an α-glucosidehydrolase inhibitor (e.g., acarbose), an α-glucosidase inhibitor (e.g.,adiposine, camiglibose, emiglitate, miglitol, voglibose, pradimicin-Q,and salbostatin), a PPARγ agonist (e.g., balaglitazone, ciglitazone,darglitazone, englitazone, isaglitazone, pioglitazone, rosiglitazone andtroglitazone), a PPAR α/γ agonist (e.g., CLX-0940, GW-1536, GW-1929,GW-2433, KRP-297, L-796449, LR-90, MK-0767 and SB-219994), a biguanide(e.g., metformin), a glucagon-like peptide 1 (GLP-1) agonist (e.g.,Byetta™, exendin-3 and exendin-4), a protein tyrosine phosphatase-1B(PTP-1B) inhibitor (e.g., trodusquemine, hyrtiosal extract, andcompounds disclosed by Zhang, S., et al., Drug Discovery Today,12(9/10), 373-381 (2007)), SIRT-1 inhibitor (e.g., reservatrol), adipeptidyl peptidease IV (DPP-IV) inhibitor (e.g., sitagliptin,vildagliptin, alogliptin and saxagliptin), an insulin secreatagogue, afatty acid oxidation inhibitor, an A2 antagonist, a c-jun amino-terminalkinase (JNK) inhibitor, insulin, an insulin mimetic, a glycogenphosphorylase inhibitor, a VPAC2 receptor agonist and a glucokinaseactivator. Preferred anti-diabetic agents are metformin, a glucagon-likepeptide 1 (GLP-1) agonist (e.g, Byetta™) and DPP-IV inhibitors (e.g.,sitagliptin, vildagliptin, alogliptin and saxagliptin).

All of the recited U.S. patents and publications, including alltechnical bulletins referenced in the Examples, are incorporated hereinby reference in their entireties.

The Examples set forth herein below are for illustrative purposes only.The compositions, methods, and various parameters reflected herein areintended only to exemplify various aspects and embodiments of theinvention, and are not intended to limit the scope of the claimedinvention in any way.

EXAMPLES

The compounds and intermediates described below were generally namedaccording to the IUPAC (International Union for Pure and AppliedChemistry) recommendations on Nomenclature of Organic Chemistry and theCAS Index rules. Unless noted otherwise, all reactants were obtainedcommercially.

Flash chromatography was performed according to the method described byStill et al., J. Org. Chem., 1978, 43, 2923.

All Biotage® purifications, discussed herein, were performed usingeither a 40M or 40S Biotage® column containing KP-SIL silica (40-63 μM,60 Angstroms) (Biotage AB; Uppsala, Sweden).

All CombiFlash® purifications, discussed herein, were performed using aCombiFlash® Companion system (Teledyne Isco; Lincoln, Nebr.) utilizingpacked RediSep® silica columns

Mass Spectra were recorded on a Waters (Waters Corp.; Milford, Mass.)Micromass Platform II spectrometer. Unless otherwise specified, massspectra were recorded on a Waters (Milford, Mass.) Micromass Platform IIspectrometer.

Proton NMR chemical shifts are given in parts per million downfield fromtetramethylsilane and were recorded on a Varian Unity 400 or 500 MHz(megaHertz) spectrometer (Varian Inc.; Palo Alto, Calif.). NMR chemicalshifts are given in parts per million downfield from tetramethylsilane(for proton) or fluorotrichloromethane (for fluorine).

The preparations described below were used in the synthesis of compoundsexemplified in the following examples.

Preparation of Starting Materials and Intermediates Carboxylic AcidStarting Materials

The following commercially available carboxylic acids were used toprepare exemplified compounds of the present invention:4-chloro-3-methylbenzoic acid (Alfa Aesar, Ward Hill, Mass.),1H-pyrazolo[3,4-b]pyridine-5-carboxylic acid (Sphinx ScientificLaboratory Product List), 1-methyl-1H-indazole-6-carboxylic acid(PharmaBlock R & D Product List), 1H-benzimidazole-5-carboxylic acid(Affinitis Pharma LLC, New Haven, Conn.), 1H-indazole-5-carboxylic acid(Tyger Scientific, Inc., Ewing, N.J.),4-amino-2-methylpyrimidine-5-carboxylic acid (Tyger Scientific, Inc.,Ewing, N.J.), 2-(methylamino)isonicotinic acid (Aurora Building Blocks),1H-pyrrolo[3,2-b]pyridine-6-carboxylic acid (Matrix Scientific),2-methyl-1H-benzimidazole-5-carboxylic acid (Apollo ScientificIntermediates for Research and Development),7H-pyrrolo[2,3-b]pyridine-2-carboxylic acid (Ryan Scientific ProductList), 1H-pyrrolo[2,3-b]pyridine-5-carboxylic acid (Matrix Scientific),2-oxoindoline-5-carboxylic acid (Apollo Scientific Intermediates forResearch and Development),2-oxo-2,3-dihydro-1H-benzimidazole-5-carboxylic acid (AKos BuildingBlocks Product List), 2-oxo-1,2,3,4-tetrahydroquinoline-7-carboxylicacid (AKos Building Blocks Product List),2-amino-1,6-naphthyridine-3-carboxylic acid (ACES Pharma Product List),3-aminoquinoxaline-2-carboxylic acid (AsisChem Screening Library),7-aminopyrazolo[1,5-a]pyrimidine-6-carboxylic acid (Ryan ScientificProduct List), 1-methyl-2-oxo-2,3-dihydro-1H-benzimidazole-5-carboxylicacid (AKos Building Blocks Product List), 4-(1H-imidazol-2-yl)benzoicacid (Sphinx Scientific Laboratory Product List),3-(1H-imidazol-4-yl)benzoic acid (Apollo Scientific Intermediates forResearch and Development),5-amino-2-phenyl-2H-1,2,3-triazole-4-carboxylic acid (Ryan ScientificScreening Library), 8-methyl-4-oxo-1,4-dihydroquinoline-2-carboxylicacid (Aurora Building Blocks), 2-carbamoylnicotinic acid (J & KScientific Product List), 8-methylimidazo[1,2-a]pyridine-2-carboxylicacid (Aurora Building Blocks), 3-(1H-pyrazol-3-yl)benzoic acid(Maybridge. Cornwall, UK), 3-(1H-pyrazol-1-yl)benzoic acid (AKosScreening Library), 1H-pyrrolo[2,3-b]pyridine-3-carboxylic acid(Aldrich), 6-morpholin-4-ylnicotinic acid (Ryan Scientific ProductList), 7-methylimidazo[1,2-a]pyridine-2-carboxylic acid (Aurora BuildingBlocks), imidazo[1,2-a]pyridine-2-carboxylic acid (Aurora BuildingBlocks), 5-pyridin-3-yl-1H-pyrazole-3-carboxylic acid (AKos ScreeningLibrary), 6-methyl-2-(methylamino)nicotinic acid (Aurora BuildingBlocks), imidazo[1,5-a]pyridine-7-carboxylic acid (Bepharm ProductList), 3H-imidazo[4,5-b]pyridine-6-carboxylic acid (Sphinx ScientificLaboratory Product List),7-hydroxypyrazolo[1,5-a]pyrimidine-6-carboxylic acid (Butt ParkScreening Library), indolizine-2-carboxylic acid (Ryan ScientificProduct List), 2-pyridin-2-yl-1H-imidazole-5-carboxylic acid (AmbinterStock Screening Collection), 3-(1H-imidazol-2-yl)benzoic acid (GreenchemInstitute Product List), pyrrolo[1,2-c]pyrimidine-3-carboxylic acid(Milestone PharmTech Product List),1H-pyrrolo[3,2-b]pyridine-2-carboxylic acid (Azasynth Building Blocks),1H-pyrrolo[3,2-c]pyridine-2-carboxylic acid (Aurora Building Blocks),imidazo[1,2-a]pyridine-7-carboxylic acid (Bepharm Product List),4-(1H-1,2,4-triazol-1-yl)benzoic acid (AKos Building Blocks ProductList), 1-methyl-1H-benzimidazole-5-carboxylic acid (AKos Building BlocksProduct List), 6-(1H-pyrazol-1-yl)nicotinic acid (Butt Park ScreeningLibrary), 1,6-naphthyridine-2-carboxylic acid (Bepharm Product List),1H-imidazo[4,5-b]pyridine-5-carboxylic acid (Sphinx ScientificLaboratory Product List),1-methyl-4-oxo-4,7-dihydro-1H-pyrazolo[3,4-b]pyridine-5-carboxylic acid(Aurora Screening Library), imidazo[1,2-a]pyridine-6-carboxylic acid(Apollo Scientific Intermediates for Research and Development),1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid (Parkway Scientific ProductList), 1H-indazole-6-carboxylic acid (Aldrich) quinoxaline-2-carboxylicacid (Aldrich), 3-acetamidobenzoic acid (Apollo Scientific Intermediatesfor Research and Development), 4-chloro-1H-indazole-6-carboxylic acid(Sinova Product List), 2-morpholinopyrimidine-5-carboxylic acid (AKosScreening Library), 1H-imidazo[1,2-b]pyrazole-6-carboxylic acid (AuroraBuilding Blocks), 3-hydroxyquinoline-4-carboxylic acid (AKos ScreeningLibrary), 8-hydroxyquinoline-7-carboxylic acid (TCI LaboratoryChemicals) and 3-(1H-pyrazol-4-yl)benzoic acid (AKos Building BlocksProduct List).

The following carboxylic acids (which were used to prepare compoundsdescribed in the Examples below) were prepared by previously publishedmeans: 3-hydroxy-6-methylpicolinic acid (P. Korovchenko et al.,Catalysis Today 2007, 121, 13-21);4-hydroxy-1,3-dimethyl-1H-pyrazole-5-carboxylic acid (Tet. Let. 1971,19, 1591); 3-amino-2,6-dimethylisonicotinic acid (Gulland, J. M.,Robinson, R. J. Chem. Soc., Trans. 1925, 127, 1493-503);5-hydroxyquinoline-6-carboxylic acid (Bogert, M. T.; Fisher, Harry L.Orig. Corn. 8th Intern. Cangr. Appl. Chem. 1912, 6, 37-44;5-hydroxyisoquinoline-6-carboxylic acid (can be prepared by hydrolysisof the corresponding methyl ester: Dyke, S. F.; White, A. W. C.;Hartley, D. Tetrahedron 1973, 29, 857-62);3-methyl-1-(pyridin-3-yl)-1H-pyrazole-5-carboxylic acid (can be preparedby analogous chemistry to J. Het. Chem. 1999, 36, 217).

The following carboxylic acid starting materials (which were used toprepare compounds described in the Examples below) were prepared asdescribed below.

Acid Preparation 1 4-chloro-1H-benzimidazole-6-carboxylic acid

To a mixture of 4-amino-3-nitrobenzoic acid (10 g, 56 mmol) in aceticacid (100 mL) at 0° C. was added sulfuryl chloride (8.98 g, 66 mmol).The reaction mixture was allowed to warm to ambient temperature andstirred overnight. The mixture was poured into ice water, filtered andair dried to give 4-amino-5-chloro-3-nitrobenzoic (7.35 g, 62%) asyellow solid. ¹H NMR (400 MHz, DMSO-d₆) δ ppm 8.24 (s, 2H) 8.19 (s, 1H)7.86 (s, 1H)

A suspension of 4-amino-5-chloro-3-nitrobenzoic (7.35 g, 34 mmol) inmethanol (150 mL) was treated with concentrated sulphuric acid (40 mL).The suspension was heated to reflux overnight. The reaction solution wasconcentrated in vacuo to give a yellow solid which was taken up in ethylacetate (200 mL) and water (30 mL). The solution was cooled to 0° C. andpotassium carbonate was added (12.4 g) in water (30 mL). The layers wereseparated and the aqueous layer was extracted with ethyl acetate (200mL). The combined organic layers were dried over magnesium sulphate andconcentrated in vacuo to give methyl 4-amino-5-chloro-3-nitrobenzoate asa yellow solid (7.25 g, 93%). ¹H NMR (400 MHz, DMSO-d₆) δ ppm 8.50 (d,J=2.15 Hz, 1H) 8.02 (d, J=1.95 Hz, 1H) 7.84 (br. s., 2H) 3.80 (s, 3H).

To a solution of methyl 4-amino-5-chloro-3-nitrobenzoate (4.29 g, 18.6mmol) in ethanol (115 mL), water (250 mL) and tetrahydrofuran (200 mL)was added sodium hydrosulfite (80 g, 391 mmol). The reaction was stirredat ambient temperature for two hours. To the reaction was added water(55 mL). After stirring for an additional hour saturated aqueous sodiumbicarbonate (140 mL) was added to the reaction. The reaction mixture wasfiltered and the filtrate was extracted twice with ethyl acetate (200 mLeach). The organic extracts were combined and washed with saturatedaqueous sodium bicarbonate (100 mL) followed by saturated aqueous sodiumchloride (100 mL). The organic layer was concentrated in vacuo to afinal volume of 100 mL and then allowed to sit at ambient temperatureovernight to give a precipitate. The mixture was filtered and driedunder a stream of nitrogen to give methyl 3,4-diamino-5-chlorobenzoate(973 mg, 26%). The filtrate was concentrated in vacuo to give methyl3,4-diamino-5-chlorobenzoate (2.45 g, 66%). ¹H NMR (400 MHz, DMSO-d₆) δppm 7.11 (d, J=1.95 Hz, 1H) 7.08 (d, J=1.95 Hz, 1H) 5.44 (s, 2H) 5.08(s, 2H) 3.70 (s, 3H).

3,4-diamino-5-chlorobenzoate (1.2 g, 6 mmol) was added to water (10 mL)and formic acid (826 mg, 18 mmol) and heated at reflux for 4 hours. Thereaction was cooled to ambient temperature and aqueous potassiumhydroxide was added (21 mL, 1M). The reaction solution was washed withethyl acetate (2×25 mL each). The aqueous layer was acidified to pH=5with aqueous hydrochloric acid (1N) to give a precipitate which wasfiltered, washed with water and dried under a stream of nitrogen to givethe title compound (439 mg, 37%). ¹H NMR (400 MHz, DMSO-d₆) δ ppm 8.45(s, 1H) 8.10 (d, J=1.17 Hz, 1H) 7.77 (d, J=1.37 Hz, 1H).

Acid Preparation 27-chloro-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylic acid

3,4-diamino-5-chlorobenzoate (from acid preparation 1, 100 mg, 0.50mmol) and carbonyl diimidazole (89 mg, 0.55 mmol) were combined intetrahydrofuran (2 mL) and stirred for 16 hours. The reaction solutionwas heated to 60° C. for 3 hours. To the reaction was added carbonyldiimidazole (81 mg, 0.50 mmol) and the reaction was continued at 60° C.for two hours. The reaction was allowed to cool to room temperature andstirred for 16 hours. A precipitate formed. The mixture was filtered.The filtrate was concentrated in vacuo and the residue was slurried inethyl acetate. The slurry was filtered on the same filter as theoriginal filtration. The collected solids were washed with a portion onethyl acetate and then dried under a stream of nitrogen to give methyl7-chloro-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylate as a whitesolid (93 mg, 82%). ¹H NMR (400 MHz, DMSO-d₆) δ ppm 11.60 (br. s., 1H)11.16 (s, 1H) 7.55 (d, J=1.37 Hz, 1H) 7.38 (d, J=1.56 Hz, 1H) 3.80 (s,3H).

Methyl 7-chloro-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylate(351 mg, 1.55 mmol), 1 M aqueous lithium hydroxide (0.774 mL, 1.55 mmol)and tetrahydrofuran (5 mL) were combined and heated to 50° C. for 2hours. To the reaction was added 1 M aqueous lithium hydroxide (0.774mL, 1.55 mmol) and methanol (10 mL) and the reaction was heated toreflux for 6 hours and then allowed to cool to ambient temperatureovernight. The reaction solution was concentrated in vacuo to remove thetetrahydrofuran and methanol. The residual aqueous layer was extractedwith ethyl acetate (2 mL). To the aqueous layer was added water (2 mL)ethyl acetate (2 mL) and 3 M aqueous hydrochloric acid. A precipitateformed. The mixture was filtered and the solids were washed with waterand ethyl acetate. The solids were dried under a stream of nitrogen togive the title compound (296 mg, 90%). ¹H NMR (400 MHz, DMSO-d₆) δ ppm11.54 (s, 1H) 11.12 (s, 1H) 7.54 (d, J=1.37 Hz, 1H) 7.38 (d, J=1.37 Hz,1H).

Acid Preparation 3 4-fluoro-1H-benzo[d]imidazole-6-carboxylic acid

To a 2.5-5 mL microwave tube, was added6-bromo-4-fluoro-1H-benzo[d]imidazole (160 mg, 0.744 mmol) suspended inde-gassed 1,4 dioxane (1.5 mL). To this was addedtrans-di(u-acetato)bis[o-(di-o-tolylphosphino)benzyl]di-palladium (II)(26 mg, 0.043 mmol) and molybdenum hexacarbonyl (100 mg, 0.38 mmol),along with sodium carbonate (237 mg, 2.23 mmol) dissolved in de-gassedwater (2 mL). The mixture was stirred for 20 seconds and then heated at155° C. in the microwave for 10 minutes, keeping the pressure under 16bar. The vessel was vented before handling and left to stand overnightat room temperature. Water (2 mL) and ethyl acetate (3 mL) were added tothe reaction, and then the mixture was filtered through Celite®. Thefiltrate was partitioned with ethyl acetate and separated. The aqueousfraction was washed with ethyl acetate once more and the combinedorganic layers were set aside. Another portion of water (5 mL) was addedto the aqueous layer and acidified with 0.5 M HCl to pH 3; a brownprecipitate was formed. The mixture was allowed to stand in therefrigerator at 4° C. for 1 hour. The mixture was filtered and washedwith water to give 4-fluoro-1H-benzo[d]imidazole-6-carboxylic acid as agrey solid, (63% yield). ¹H NMR (500 MHz, DMSO-d₆) □ ppm 12.99 (br. s.,1H) 8.47 (s, 1H) 8.02 (s, 1H) 7.52 (d, J=11.71 Hz, 1H).

Acid Preparation 4 1-oxo-1,2-dihydroisoquinoline-6-carboxylic acid

To a mixture of (E)-3-(3-bromophenyl)acrylic acid (100 g, 0.44 mol) andtriethylamine (0.48 mol) in toluene (1000 mL) was addeddiphenylphosphoryl azide (127.4 g, 0.45 mol) dropwise at 0-10° C. Themixture was stirred at room temperature overnight. Thin layerchromatography (petroleum ether/ethyl acetate=8:1) indicated completionof reaction. The resulting mixture was washed with 1 N sodium hydroxide(500 mL) and extracted with ethyl acetate (2000 mL×3). The organic layerwas concentrated to give crude(E)-1-azido-3-(3-bromophenyl)prop-2-en-1-one, which was used in the nextstep directly.

A mixture of crude (E)-1-azido-3-(3-bromophenyl)prop-2-en-1-one (crudeabout 120 g) and toluene (200 mL) was refluxed for two hours. Thin layerchromatography (petroleum ether/ethyl acetate=8:1) indicated most of thestarting material was consumed. The mixture was concentrated to givecrude (E)-1-bromo-3-(2-isocyanatovinyl)benzene (100 g, 94%), which wasused in the next step directly.

A solution of (E)-1-bromo-3-(2-isocyanatovinyl)benzene (100 g, 0.44 mol)in toluene (200 mL) was added dropwise to a mixture of tributylamine(100 mL) and oxydibenzene (500 mL) at 190° C. After the addition, themixture was heated at 210° C. for another two hours. Thin layerchromatography (petroleum ether/ethyl acetate=1:1) indicated thereaction was complete. The mixture was cooled to room temperature,filtered, and the solid was washed with ethyl acetate (50 mL×3). Thesolid was dried under vacuum to give crude 6-bromoisoquinolin-1(2H)-one(30 g, 30%) as a light yellow solid, which was used in the next stepdirectly.

A mixture of 6-bromoisoquinolin-1(2H)-one (30 g, 134 mmol),triethylamine (17.6 g, 174 mmol), palladium (II) chloride (0.24 g, 1.34mmol) and (S)-(−)-2,2′-bis(diphenylphosphino)-1,1′-binaphthyl (0.84 g,1.34 mmol) in methane (300 mL) was heated at 100° C. under 2 MPa ofcarbon monoxide and stirred for 12 hours. Thin layer chromatography(petroleum ether/ethyl acetate=1:1) indicated the reaction was complete.The reaction mixture was concentrated, the residue was washed withwater, and the solid was filtered and dried in vacuum to give crudemethyl 1-oxo-1,2-dihydroisoquinoline-6-carboxylate (23.8 g, 95%) as ayellow solid, which was used in the next step directly.

To a mixture of methyl 1-oxo-1,2-dihydroisoquinoline-6-carboxylate (25g, 0.133 mol), tetrahydrofuran (200 mL) and water (200 mL) was addedlithium hydroxide (16.8 g, 0.40 mol) at room temperature, and themixture was stirred for four hours. Thin layer chromatography (petroleumether/ethyl acetate=1:1) indicated the reaction was complete. Thereaction mixture was extracted with ethyl acetate (100 mL×3) to removeimpurities. The aqueous layer was acidified with 4 N aqueous HCl to pH 5and filtered. The solid was dried in vacuum to give1-oxo-1,2-dihydroisoquinoline-6-carboxylic acid (11.3 g, 48%) as a lightyellow solid. ¹H NMR (400 MHz, DMSO-d₆) δ ppm 11.48 (s, 1H), 8.24 (d,2H), 7.93 (d, 1H), 7.22 (d, 1H), 6.68 (d, 1H).

Acid Preparation 5 1-oxo-1,2-dihydroisoquinoline-7-carboxylic acid

1-oxo-1,2-dihydroisoquinoline-7-carboxylic acid was prepared in ananalogous fashion to 1-oxo-1,2-dihydroisoquinoline-6-carboxylic acid,(acid preparation 4).

Acid Preparation 6 5-(1H-imidazol-1-yl)picolinic acid

5-bromopicolinonitrile (2.0 g, 10.9 mmol), imidazole (818 mg, 12 mmol)potassium carbonate (1.66 g, 12 mmol) and dimethylformamide (40 mL) werecombined and heated to 130° C. for 20 hours. The reaction solution wasevaporated and the residue was partitioned between dichloromethane (150mL) and water (100 mL). The phases were separated and the organic phasewas washed with water (50 mL), dried over magnesium sulfate andevaporated to give a residue which was purified by flash chromatographyeluting with 2-3% methanol in dichloromethane gradient to give5-(1H-imidazol-1-yl)picolinonitrile (1.23 g, 66%).

5-(1H-imidazol-1-yl)picolinonitrile (136 mg, 0.80 mmol) was heated toreflux in 6N aqueous hydrochloric acid (10 mL) for 2 hours. The reactionmixture was evaporated and the residue was azeotroped with threeportions of toluene to give a residue which was purified on an ionexchange column (AG-50 Biorad) eluting with a 0-10% pyridine in watergradient to give the title compound as a white solid (128 mg, 84%). ¹HNMR (400 MHz, DMSO-d₆) δ ppm 9.10 (s, 1H), 8.50 (s, 1H), 8.26-8.33 (m,1H), 8.13-8.20 (m, 1H), 7.96 (s, 1H), 7.16 (s, 1H).

Acid Preparation 7 7-chloro-1H-indazole-5-carboxylic acid

To a mixture of 4-amino-3-chloro-5-methyl-benzonitrile (3.0 g, 18.0mmol) in chloroform (50 mL) was added acetic anhydride (3.92 mL, 41.4mmol). The mixture was heated at reflux for 5 hours and then cooled toroom temperature. To the mixture was added potassium acetate (530 mg,5.4 mmol) and isoamyl nitrite (5.28 mL, 39.6 mmol). The reaction washeated at reflux for 16 hours. The reaction mixture was cooled to roomtemperature, extracted with saturated aqueous sodium bicarbonate, theorganics were dried over sodium sulfate, and concentrated in vacuo toafford a brown oil. The oil was dissolved in methanol (25 mL) andconcentrated hydrochloric acid (25 mL) was added. The reaction wasstirred at room temperature for 22 hours and the methanol wasconcentrated in vacuo. The remaining aqueous layer was adjusted to a pHof 7 and the resultant precipitate was filtered to afford a brown solidwhich was purified by flash chromatography using 50% dichloromethane inheptane as eluent to afford 7-chloro-1H-indazole-5-carbonitrile as asolid (585 mg, 18%): −ESI MS (M−1) 176.0; ¹H NMR (400 MHz, CDCl₃) δ ppm8.29 (br. s., 2H), 8.08 (s, 1H), 7.61 (s, 1H).

To a mixture of 7-chloro-1H-indazole-5-carbonitrile (1.36 g, 7.66 mmol)in ethanol (52.5 mL) was added water (17.5 mL) and potassium hydroxide(6.44 g, 115 mmol). The reaction mixture was heated at reflux for 16hours. The reaction mixture was cooled to room temperature, extractedtwice with ethyl ether, acidified the aqueous with 1N hydrochloric acidand the resultant precipitate was filtered to afford7-chloro-1H-indazole-5-carboxylic acid as a brown solid (900 mg, 60%):−ESI MS (M−H) 195.2.

Acid Preparation 8 5-morpholinopicolinic acid

Diethyl malonate (151 g, 0.944 mol) was added dropwise under stirring to60% sodium hydride in mineral oil (37.8 g, 0.944 mol) in drytetrahydrofuran (1 L). After hydrogen evolution ceased,2-chloro-5-nitropyridine (125 g, 0.787 mol) was added. The reactionmixture was refluxed for 2 hours and then the tetrahydrofuran wasevaporated in vacuo to give crude diethyl (5-nitropyridin-2-yl)malonate,which was used at the next stage without purification.

Crude diethyl (5-nitropyridin-2-yl)malonate was added to boiling 65%nitric acid (1.5 L) under stirring. The reaction mixture was refluxedunder stirring for 15 hours. The reaction mixture was concentrated invacuo and the resulting solid was washed with chloroform to give5-nitropyridine-2-carboxylic acid (yield 65%, 85.9 g).

5-Nitropyridine-2-carboxylic acid (100 g, 0.60 mol) was heated at refluxin methanol (1 L) and sulfuric acid (57 mL) for 5 hours. The reactionmixture was cooled, reduced to half volume in vacuo, and the residueneutralized with a solution of sodium carbonate. The resultingprecipitate was filtered to give methyl 5-nitropyridine-2-carboxylate(yield 89%, 98

Methyl 5-nitropyridine-2-carboxylate (182 g, 1 mol) was refluxed inpiperidine (250 mL) for 1 hour. The reaction mixture was concentrated invacuo to give crude 5-nitro-2-(piperidin-1-ylcarbonyl)pyridine, whichwas used for the next stage without additional purification.

Crude 5-nitro-2-(piperidin-1-ylcarbonyl)pyridine was reduced by hydrogenunder atmospheric pressure in the presence of 10% palladium on carbon (4g) in acetic acid (500 mL). The catalyst was separated by filtration andthe solvent evaporated in vacuum to give crude6-(piperidin-1-ylcarbonyl)pyridin-3-amine, which was used for the nextstage without additional purification.

A solution of sodium nitrite (69 g) in concentrated hydrochloric acid(1.5 L) was added to crude 6-(piperidin-1-ylcarbonyl)pyridin-3-amine at0 C, and the mixture was stirred for 10 minutes. Urea (20 g) was added,and the mixture was stirred for 15 minutes. Sodium iodide (150 g) wasadded, and the product was separated by filtration and recrystallizedfrom ethanol to give 5-iodo-2-(piperidin-1-ylcarbonyl)pyridine (yield23% calculated for methyl 5-nitropyridine-2-carboxylate, 71 g).

A mixture of 5-iodo-2-(piperidin-1-ylcarbonyl)pyridine (71 g, 0.23 mol),palladium (II) acetate (1.03 g, 46 mmol),2-(di-tert-butylphosphino)biphenyl (2.76 g, 92 mmol), morpholine (23.7g, 0.28 mol), and sodium tert-butoxide (27.8 g, 0.28 mol) in toluene(400 mL) was stirred under argon at 95 C for 2 hours. The product wasisolated by chromatography (silica gel, ethyl acetate) andrecrystallized from ethanol to give4-[6-(piperidin-1-ylcarbonyl)pyridin-3-yl]morpholine (yield 37%, 22 g).

25% KOH (100 mL) was added to4-[6-(piperidin-1-ylcarbonyl)pyridin-3-yl]morpholine (18.3 g), and themixture was refluxed and then neutralized with HCl. The solution wasevaporated in vacuum, and the product was extracted with hot isopropanolto give the title compound (yield 71%, 11.5 g). +ESI MS (M+H) 209.7; ¹HNMR (400 MHz, DMSO-d₆) δ ppm 8.34 (br. s., 1H), 8.03 (d, 1H), 7.65 (dd,1H), 3.75 (br. s., 4H), 3.40 (br. s., 4H).

Acid Preparation 9 7-chloro-2-methyl-1H-benzo[d]imidazole-5-carboxylicacid

Add 2N hydrochloric acid (8 mL) to a solution of3,4-diamino-5-chlorobenzoate (from acid preparation 1, 435 mg, 2.17mmol) in ethanol (20 mL). Heat the mixture to reflux then addacetylacetone (437 mg, 4.37 mmol) to the yellow solution. The yellowsolution turned purple upon addition. Stir at reflux for 1 hour and thesolution turned back to yellow. Stir at reflux for an additional 1 hour.Concentrate the solvent to a colorless residue. Add water (20 mL).Extract the suspension with ethyl acetate (20 mL). Basify the aqueouslayer with 2N sodium hydroxide (˜8 mL) to pH˜10. Extract with ethylacetate (3×15 mL). Wash combined organics from the basic extraction withbrine (5 mL). Dry over magnesium sulfate, filter, concentrate, and dryunder high vacuum to yield methyl7-chloro-2-methyl-1H-benzo[d]imidazole-5-carboxylate (290 mg, 59%) as acolorless solid. ¹H NMR (400 MHz, CDCl₃) δ ppm 2.68 (s, 3H), 3.93 (s,3H), 7.25 (s, 1H), 7.96 (s, 1H).

Add 2N sodium hydroxide (5 mL, 5 mmol) to a solution of methyl7-chloro-2-methyl-1H-benzo[d]imidazole-5-carboxylate (280 mg, 1.25 mmol)in methanol (7.5 mL). Stir at 65° C. for 16 hours. The methanol wasconcentrated in vacuo and the remaining aqueous layer was extracted withethyl acetate (10 mL). Acidify the aqueous layer to pH˜4 with 1Nhydrochloric acid (˜5 mL). A colorless precipitate was filtered anddried under high vacuum to yield the title compound (189 mg, 72%). ¹HNMR (400 MHz, CD₃OD) δ ppm 2.61 (s, 3H), 7.86 (d, J=1.37 Hz, 1H), 8.08(d, J=1.17 Hz, 1H).

Acid Preparation 10 7-chloro-2-methyl-1H-benzo[d]imidazole-5-carboxylicacid

A round bottomed flask was charged with5-bromo-3-fluorobenzene-1,2-diamine (400 mg, 2 mmol) and 30 mL ethanol.5 N hydrochloric acid (8 mL, 40 mmol) was then added. This mixture washeated to reflux and 2,4-pentanedione was added. The reaction mixtureturned deep purple in color then slowly turned back to tan. Reaction wasallowed to proceed for 3 hours and then cooled and neutralized withsaturated sodium bicarbonate solution. The reaction mixture was thenextracted three times with dichloromethane. The combined organic layerswere washed with brine, dried with magnesium sulfate, filtered andconcentrated in vacuo. The crude mixture was triturated in diethyl etherthen filtered to give 6-bromo-4-fluoro-2-methyl-1H-benzo[d]imidazole(375 mg, 82%) as a tan solid. ¹H NMR (400 MHz, CDCl₃) δ ppm 7.44 (br.s., 1H), 7.10 (d, J=11.22 Hz, 1H), 2.63 (s, 3H).

A 5 mL microwave vial was charged6-bromo-4-fluoro-2-methyl-1H-benzo[d]imidazole (187 mg, 0.815 mmol) andsuspended in de-gassed dioxane (2 mL),trans-di-μ-acetatobis[2-(di-O-tolylphosphino)benzyl]dipalladium (II) (28mg, 0.048 mmol) and molybdenumhexacarbonyl (110 mg, 0.417 mmol).Degassed 10% aqueous sodium carbonate (2.45 mL, 2.45 mmol) was thenadded. The reaction was then stirred for 20 seconds before being reactedin the microwave at 155° C. at very high absorption for 10 minutes. Thevessel was then vented and left to stand overnight at room temperature.Water (2 mL) and ethyl acetate (3 mL) were then added and the mixturewas filtered through Celite®. The layers were separated and the aqueouslayer was washed with ethyl acetate (×2). The combined ethyl acetatelayers were set aside. Water (5 mL) was added to the aqueous layer whichwas then acidified with 0.5 M hydrochloric acid to a pH of 3 then cooledto 4° C. A solid formed which was filtered and washed with water to givethe title compound (61 mg, 37%) as a yellow solid. A second crop formedwhich was then filtered to give the title compound (100 mg, 63%). ¹H NMR(500 MHz, CD₃OD) δ ppm 8.22 (d, J=0.98 Hz, 1H), 7.91 (d, J=10.49 Hz,1H), 2.95 (s, 3H).

Acid Preparation 11 1H-pyrazolo[4,3-b]pyridine-6-carboxylic acid

To a suspension of sodium hydride (5.08 g, 127 mmol) indimethylformamide (75 mL) was added diethyl malonate (19.26 mL, 127mmol) at 0° C. The solution was then stirred at ambient temperature for30 minutes and a solution of 5-bromo-2-chloro-3-nitropyridine (30 g, 127mmol) in dimethylformamide (75 mL) was added dropwise. The dark brownmixture was then stirred at 100° C. for 2 hours before being cooled toambient temperature and quenched with a saturated solution of ammoniumchloride (500 mL) at 0° C. The mixture was extracted with ethyl acetate(3×500 mL) and the combined organics were dried over magnesium sulfate.The solvent was removed in vacuo to give a dark brown oil which waspurified by flash column chromatography (10% ethyl acetate/hexane) toafford diethyl 2-(5-bromo-3-nitropyridin-2-yl)malonate as a brown solid(31.8 g, 88 mmol, 69%). ¹HNMR (400 MHz, CDCl₃): δ ppm 8.86 (s, 1H), 8.61(s, 1H), 5.44 (1H, s), 4.29 (q, 4H), 1.27 (t, 6H).

A mixture of the diethyl 2-(5-bromo-3-nitropyridin-2-yl)malonate (31.8g, 88 mmol) in aqueous hydrochloric acid (6M, 1.4 L) was stirred atreflux for 18 hours. The reaction mixture was cooled to ambienttemperature and added very slowly to a saturated aqueous solution ofaqueous sodium bicarbonate (4 L) at 0° C. The mixture was then extractedwith dichloromethane (7 L), dried over magnesium sulfate and the solventremoved in vacuo to give 5-bromo-2-methyl-3-nitropyridine as an orangeoil (13.8 g, 63.9 mmol, 72%) which solidified upon standing. ¹HNMR (300MHz, CDCl₃): δ ppm 8.78 (s, 1H), 8.43 (s, 1H), 2.79 (s, 3H).

To a solution of 5-bromo-2-methyl-3-nitropyridine (13.8 g, 63.9 mmol) inindustrial methylated spirit (330 mL) at 40° C. was added iron powder(20 g) (portionwise to avoid clumping) followed by concentrated aqueoushydrochloric acid (5 mL). The dark brown mixture was stirred vigorouslyat reflux for 2 hours and then cooled and filtered through Celite®(which was washed with 1 L of industrial methylated spirit). The solventwas then removed in vacuo and the residue taken up in ethyl acetate (200mL) and washed with a saturated aqueous solution of sodium bicarbonate(200 mL), dried over magnesium sulfate and solvent removed in vacuo togive 5-bromo-2-methylpyridin-3-amine as an orange solid, (10.7 g, 57.5mmol, 89.9%). ¹HNMR (400 MHz, CDCl₃): δ ppm 7.91 (s, 1H), 7.00 (s, 1H),3.75 (br.s, 2H), 2.25 (s, 3H).

To a solution of 5-bromo-2-methylpyridin-3-amine (10.7 g, 57.5 mmol) indichloromethane (575 mL) was added acetic anhydride (12 mL, 126.5 mmol)at 0° C. followed by triethylamine (22 mL, 158 mmol). The mixture wasallowed to warm to ambient temperature and stirred for 18 hours at whichpoint a further equivalent of acetic anhydride (6 mL, 63 mmol) wasadded. The mixture was stirred at ambient temperature for a further 72hours. The reaction mixture was quenched with a saturated aqueoussolution of sodium bicarbonate (500 mL) and the organic phase washedwith saturated aqueous sodium chloride (500 mL), dried over magnesiumsulfate and concentrated in vacuo to give a brown solid. This solid wastriturated with 30% ethyl acetate in hexanes to yieldN-(5-bromo-2-methylpyridin-3-yl)acetamide as an off-white solid, (8.28g, 36 mmol, 63%). ¹HNMR (400 MHz, CD₃OD): δ ppm 8.31 (s, 1H), 8.18 (s,1H), 2.43 (s, 3H), 2.18 (s, 3H).

To a solution of N-(5-bromo-2-methylpyridin-3-yl)acetamide (8.28 g, 36mmol) in chloroform (550 mL) at ambient temperature was added potassiumacetate (4.32 g, 43.6 mmol), acetic acid (2.5 mL, 43.6 mmol) andfollowed by acetic anhydride (6.86 mL, 72.6 mmol). The mixture wasstirred at ambient temperature for 15 minutes before being heated to 40°C. Isoamylnitrite was then added dropwise. The reaction was then stirredat 60° C. for 48 hours. The reaction mixture was poured slowly into asaturated solution of sodium bicarbonate (500 mL) at 0° C. The organicphase was retained and the aqueous phase extracted with dichloromethane(500 mL). The combined organics were then concentrated to a brown oilwhich was dissolved in methanol (500 mL). Aqueous sodium hydroxide (2 M,500 mL) was added at 0° C. and the mixture stirred at ambienttemperature for 1 hour before the methanol was removed in vacuo. Theaqueous mixture was then extracted with ethyl acetate (3×500 mL). Thecombined organics dried over magnesium sulfate, and the solvent removedin vacuo to give 6-bromo-1H-pyrazolo[4,3-b]pyridine as a light brownsolid (5.5 g, 27.9 mmol, 77%). ¹HNMR (400, CD₃OD): δ ppm 8.55 (s, 1H),8.24 (s, 1H), 8.21 (s, 1H).

To a solution of 6-bromo-1H-pyrazolo[4,3-b]pyridine (5.5 g, 27.9 mmol)in methanol (125 mL) and acetonitrile (75 mL) was added triethylamine(22 mL, 156 mmol), 2,2′-bis(diphenylphosphino)-1,1′-binaphthyl (1.98 g,3.07 mmol), palladium dichloride (1.23 g, 6.98 mmol). The mixture wasplaced under 20 bar of carbon monoxide, heated to 100° C., and stirredvigorously for 18 hours. The reaction mixture was cooled to ambienttemperature and filtered through Celite® before the solvent was removedin vacuo to yield a brown oil. This crude oil was then purified by flashcolumn chromatography (1:1, ethyl acetate:hexane) to give methyl1H-pyrazolo[4,3-b]pyridine-6-carboxylate as a pale yellow solid (4.52 g,92% yield). ¹HNMR (400, CDCl₃) δ ppm 10.56 (s, 1H), 9.23 (s, 1H), 8.35(s, 1H), 8.40 (s, 1H), 4.01 (s, 3H).

To a solution of methyl 1H-pyrazolo[4,3-b]pyridine-6-carboxylate (3.52g, 20 mmol) in methanol (250 mL) and water (190 mL) at 0° C. was addedaqueous sodium hydroxide (2M, 64 mL, 128 mmol), dropwise. The suspensionwas then allowed to warm to ambient temperature and stirred for 18hours. The methanol was then removed in vacuo and the aqueous mixtureextracted with ethyl acetate (250 mL) before being acidified (to pH 5-6)with aqueous hydrochloric acid (2 M, 70 mL). The cream solid which hadprecipitated out was then filtered off and dried in a desiccator toyield the title compound (0.675 g, 4.16 mmol, 21% yield). ¹HNMR (400MHz, DMSO-d₆): δ ppm 8.97 (s, 1H), 8.45 (s, 1H), 8.39 (s, 1H).

Acid Preparation 12 3-cyano-1H-indazole-5-carboxylic acid

A suspension of (2-nitrophenyl)-acetonitrile (30 g, 185 mmol) and 10%palladium on carbon (2 g) in acetic acid (450 mL) was hydrogenated in aParr apparatus under 30 psi pressure at ambient temperature for 2 hours.The mixture was filtered through a Celite® pad and the filtrate wasconcentrated in vacuo. The obtained residue was dissolved in ethylacetate (250 mL). The resulting solution was washed with water (2×100mL) and saturated sodium chloride (50 mL), and then dried over anhydroussodium sulfate and concentrated in vacuo to yield product. The crudematerial was purified by column chromatography (100-200 mesh silica gel)using 8% ethyl acetate in petroleum ether as eluent to afford(2-aminophenyl)acetonitrile (13.5 g, 55%) as a solid. ¹HNMR (CDCl₃) δppm 7.3-7.1 (m, 2H), 6.9-6.7 (m, 2H), 3.7 (br, 2H), 3.5 (s, 2H).

To a cooled solution of (2-aminophenyl)acetonitrile (13 g, 98 mmol) indimethylformamide (150 mL) at 0° C., was added N-bromosuccinimide (19.3g, 108 mmol) in portions for 30 minutes and maintained at 0° C. for 1hour. The mixture was diluted with ethyl acetate (300 mL) and washedwith water (3×100 mL) and saturated sodium chloride (50 mL). The organiclayer was dried over anhydrous sodium sulfate and concentrated in vacuo.The obtained crude product was purified by column chromatography(100-200 mesh silica gel) using 10% ethyl acetate in petroleum ether aseluent to afford (2-amino-5-bromophenyl)acetonitrile (11 g, 53%) assolid. ¹HNMR (CDCl₃) δ 7.35 (s, 1H), 7.25 (d, 1H), 6.65 (d, 1H), 3.7(br, 2H), 3.52 (s, 2H).

To a cooled solution of (2-amino-5-bromophenyl)acetonitrile (11 g, 52mmol) in concentrated hydrochloric acid (110 mL) at −50° C., a solutionof sodium nitrite (3.9 g, 57 mmol) in water (20 mL) was added slowly.After the addition, the mixture was stirred at −50° C. for 2 h. Themixture was neutralized with 33% ammonium hydroxide at 0° C. andextracted with ethyl acetate (3×100 mL). The combined organic layerswere washed with saturated sodium chloride (100 mL), dried overanhydrous sodium sulfate and concentrated. The obtained crude productwas purified by column chromatography (100-200 mesh silica gel) using10% ethyl acetate in petroleum ether as eluent to afford5-bromo-3-cyanoindazole (7 g, 60%) as a solid. ¹HNMR (CDCl₃) δ ppm 10.7(br, 1H), 8.1 (s, 1H), 7.64 (d, 1H), 7.5 (d, 1H).

A suspension of 5-bromo-3-cyanoindazole (3 g, 13.51 mmol), palladiumdichloride 1,1′-bis(diphenylphosphino)ferrocene (1.76 g, 2.16 mmol),sodium acetate (3.32 g, 40.5 mmol), dimethylformamide (1 mL) in methanol(100 mL) was degassed, and kept under carbon monoxide (80 psi) pressureat 80° C. in a autoclave for 16 hours. The mixture was diluted withwater (50 mL), filtered through Celite® bed and the filtrate wasconcentrated. The obtained residue was acidified with 10% citric acidsolution and extracted with ethyl acetate (2×100 mL). The combinedorganic layers were washed with brine (50 mL), dried over anhydroussodium sulfate and concentrated. The obtained crude product was purifiedby column chromatography (100-200 mesh silica gel) using 10% ethylacetate in chloroform as eluent to afford methyl3-cyano-1H-indazole-5-carboxylate (1.8 g, 68%) as a solid. ¹HNMR (CDCl₃)δ ppm 10.8 (s, 1H), 8.7 (s, 1H), 8.22 (d, 1H), 7.64 (d, 1H), 4.0 (s,3H).

To a solution of methyl 3-cyano-1H-indazole-5-carboxylate (2.5 g, 12mmol) in ethanol (40 mL), a solution of lithium hydroxide (1.04 g, 24.9mmol) in water (15 mL) was added and stirred at ambient temperature for16 h. The mixture was concentrated and the obtained residue wasdissolved in water (25 mL) and washed with ethyl acetate (20 mL). Theaqueous layer was acidified with 10% citric acid solution, the obtainedprecipitate was filtered, washed with 50% ethyl acetate in petroleumether (2×10 mL) and dried to afford the title compound (1.9 g, 82%) as abrown solid. ¹HNMR (DMSO-d₆) δ ppm 13.8-12.4 (br, 2H), 8.44 (s, 1H), 8.1(d, 1H), 7.82 (d, 1H).

Acid Preparation 13 2-(1H-pyrazol-3-yl)isonicotinic acid

To a stirred solution of 29.0 g (69 mmol) 2-bromo-4-methylpyridine in150 mL concentrated sulfuric acid was added portionwise 67.9 g (231mmol) potassium dichromate. The reaction mixture was cooled with an icebath so that the temperature stayed between 20-50° C. After the additionwas complete, stirring was continued at room temperature for a further 2hours. The reaction mixture was then poured slowly onto 2 L ice-waterand the mixture stirred for 1 hour at room temperature. The resultingcrystals were collected by filtration, washed with water until thewashings were colorless, and dried in vacuo to afford 30.0 g (88%) of2-bromoisonicotinic acid.

To an ice cooled solution of 2-bromoisonicotinic acid (73 g, 0.361 mol)in dichloromethane (500 mL) and methanol (35 g, 1.08 mol) was added1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride (67 g,0.434 mol) by portions. The mixture was stirred at ambient temperatureovernight. Then 120 g silica gel was added and the solvent evaporated.The residue was purified by flash chromatography, eluting with 5% ethylacetate in petroleum ether to afford 58 g (75%) of methyl2-bromoisonicotinate as a white solid.

Methyl 2-bromoisonicotinate (216 g, 1 mol), dry acetonitrile (1.7 L),ethynyl(trimethyl)silane (117 g, 1.2 mol), diisopropylamine (122 g, 1.2mol), and dichlorobis(triphenylphosphine)palladium (36 g, 0.05 mol) wereplaced into a well dried three necked flask which was twice purged witha stream of nitrogen. The reaction mixture was stirred for 0.5 hours,cooled to 10° C., and copper iodide (19 g, 0.1 mol) was added under astream of nitrogen. At 20° C., the reaction mixture became thick andblack and an exotherm was observed which was followed by formation of aprecipitate. After the addition of copper iodide, the reaction mixturewas stirred for further 2 hours at ambient temperature. The precipitatedresidue was separated by filtration and twice washed with diethyl ether(800 mL). The filtrate was washed with saturated ammonium chloride(2×300 mL) and brine (2×300 mL). After drying over sodium sulfate, thesolvent was evaporated. The residue was purified using a silica gelcolumn, eluting with hexane followed by 5% ethyl acetate in petroleumether to yield 191 g (82%) of methyl2-((trimethylsilyl)ethynyl)isonicotinate.

Concentrated sulfuric acid (60 mL, 1.1 mol) was added to a suspension ofmethyl 2-((trimethylsilyl)ethynyl)isonicotinate (127 g, 0.54 mol) intetrahydrofuran (600 mL) and mercury acetate (51.5 g, 0.16 mol). Thesuspension was stirred for 3 hours at 50° C. and kept overnight. Thereaction mixture was diluted with diethyl ether (1.5 L) and the sulfuricacid was neutralized with saturated sodium bicarbonate (150 g, 1.7 mol).A residue of mercury salts was separated by filtration. The ethersolution was washed with water and dried over sodium sulfate. Thesolvent was removed to give methyl 2-acetylisonicotinate as an oil thatwas directly used in the next step.

To a 2 L three necked flask was added methyl 2-acetylisonicotinate (160g, 0.894 mol), dimethylformamide-dimethylacetamide (350 mL) and toluene(350 mL). The mixture was refluxed for about 5 hours with a Dean-Starktrap to remove methanol produced. Additionaldimethylformamide-dimethylacetamide and toluene was added to keep thereaction volume at about 800-900 mL. When liquid chromatography-massspectrometry showed reaction completed, the solvent was removed to yieldcrude (Z)-methyl 2-(3-(dimethylamino)acryloyl)isonicotinate as a darksolid. The crude solid was directly used in the next step.

To a 2 L three-necked flask was added (Z)-methyl2-(3-(dimethylamino)acryloyl)isonicotinate (0.894 mol), hydrazinehydrate (48.8 g), anhydrous ethanol (1 L). The suspension was stirred at20° C. overnight. The solvent was removed in vacuo. The residue wastaken up in concentrated hydrochloric acid (600 mL) and heated to refluxfor 2 hours. The mixture was cooled to ambient temperature. Theresultant precipitate was filtered, washed with water, ethanol andacetone and dried to give 78.6 g of the title compound as a brown solid.¹HNMR (DMSO-d₆/D₂O) δ ppm 8.80 (d, 1H), 8.50 (s, 1H), 7.91 (d, 1H), 7.87(dd, 1H), 7.15 (d, 1H).

Acid Preparation 143-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylic acid

Add an aqueous solution of sodium hydrosulfite (17.4 g, 100 mmol in 80mL of water) to methyl-3-(methylamino)-4-nitrobenzene carboxylate (855mg, 4.75 mmol) in tetrahydrofuran (70 mL) and ethanol (30 mL) at 0° C.The orange solution turned to an orange suspension upon addition. Stirthe mixture at room temperature for 2 hours. The orange suspensionturned to a yellow suspension over this time. Add saturated sodiumbicarbonate (100 mL) then the yellow suspension turned colorless.Extract the mixture with ethyl acetate (2×100 mL). Wash the combinedorganics with brine (30 mL). Dry over magnesium sulfate, filter,concentrate, and dry under high vacuum to yield methyl4-amino-3-(methylamino)benzoate (586 mg, 68%) as a yellow oil. Theresulting oil began to crystallize upon standing after 10 minutes. ¹HNMR (400 MHz, CDCl₃) δ ppm 2.89 (s, 3H), 3.37-3.81 (m, 2H), 3.85 (s,3H), 6.67 (d, J=8.01 Hz, 1H), 7.33 (d, J=1.37 Hz, 1H), 7.44 (dd, J=8.11,1.66 Hz, 1H).

Add carbonyl diimidazole (567 mg, 3.50 mmol) to a solution of methyl4-amino-3-(methylamino)benzoate (586 mg, 3.19 mmol) in tetrahydrofuran(20 mL) at room temperature. Stir the yellow solution at roomtemperature for 16 hours. Add carbonyl diimidazole (500 mg, 0.96). Stirat room temperature for 4 hours. Add ethyl acetate (75 mL). Wash with10% citric acid (5 mL), 1N sodium hydroxide (5 mL), and brine (5 mL).Dry the organics over magnesium sulfate, filter, concentrate, and dryunder high vacuum to yield a crude yellow solid (690 mg, 100%).Triturate this crude solid with ethyl acetate (10 mL). Filter theprecipitate and dry under high vacuum to yield methyl3-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylate (422 mg,64%). ¹H NMR (400 MHz, CDCl₃) δ ppm 3.46 (s, 3H), 3.92 (s, 3H), 7.12 (d,J=8.21 Hz, 1H), 7.68 (s, 1H), 7.84 (dd, J=8.31, 1.47 Hz, 1H), 9.87-10.03(m, 1H).

Add 1N sodium hydroxide (6.1 mL, 6.1 mmol) to a suspension of methyl3-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylate (420 mg,2.04 mmol) in methanol (10 mL). The suspension turned to a solution uponaddition of 1N sodium hydroxide. Stir at 65° C. for 16 hours. Cool toroom temperature then concentrate to remove the methanol. Extract theaqueous with ethyl acetate (5 mL). Acidify the aqueous with 2N hydrogenchloride (3 mL) to pH˜2. Concentrate the aqueous layer to a solid.Triturate the solid with water (3 mL). Filter the precipitate and dryunder high vacuum to yield the title compound (234 mg, 59%) as a palebrown solid. ¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.28 (s, 3H), 7.01 (d,J=8.21 Hz, 1H), 7.57 (s, 1H), 7.63 (dd, J=8.11, 1.27 Hz, 1H), 11.19 (s,1H), 12.60 (s, 1H).

Acid Preparation 157-bromo-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylic acid

A suspension of methyl 4-amino-3-bromo-5-nitrobenzoate (10 g, 36.3 mmol)and tin(II) chloride (33 g, 14.5 mmol) in methanol (100 mL) was heatedto 60° C. and maintained for 4 hours. The reaction mass was cooled toambient temperature and concentrated to obtain a residue; the residuewas basified using saturated aqueous sodium bicarbonate until pH was 11and the aqueous layer was extracted with dichloromethane (3×200 mL). Thecombined organic layers were washed with aqueous saturated sodiumchloride (200 mL), dried over anhydrous sodium sulfate and concentratedto obtain methyl 3,4-diamino-5-bromobenzoate as an off-white solid (5 g,58%). ¹HNMR (CDCl₃): δ ppm 7.74 (s, 1H), 7.35 (s, 1H), 4.18 (broad s,2H), 3.85 (s, 3H) and 3.38-3.56 (broad s, 2H).

A solution of 3,4-diamino-5-bromobenzoate (1 g, 4.0 mmol) andtriethylamine (0.4 g, 4.0 mmol) in dichloromethane (6 mL) was cooled to0° C. A solution of triphosgene (1.2 g, 4.08 mmol) in dichloromethane(15 mL) was added to this solution. The reaction mixture was allowed towarm to ambient temperature and maintained for 18 hours. The reactionmass was quenched with water (3 mL) and extracted with ethyl acetate(3×10 mL). The combined organic layers were washed with aqueoussaturated sodium chloride (50 mL), dried over anhydrous sodium sulfateand concentrated to obtain methyl7-bromo-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylate as anoff-white solid (500 mg, 45%). ¹HNMR (CDCl₃+DMSO-d₆): δ 11.35 (s, 1H),11.05 (s, 1H), 7.75 (s, 1H), 7.52 (s, 1H) and 3.85 (s, 3H).

Methyl 7-bromo-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylate (238mg, 0.878 mmol) and 2 N aqueous sodium hydroxide (1.50 mL, 3.0 mmol)were combined in methanol (5 mL) and heated to 50° C. for 90 minutes.The reaction solution was concentrated to remove the methanol. To thereaction residue was added ethyl acetate (5 mL). The resultant solutionwas acidified with 1N aqueous hydrochloric acid (1.5 mL) to give a finalpH of 4. A precipitate formed which was filtered and dried under vacuumto give the title compound (226 mg, 100%) as a solid. ¹H NMR (400 MHz,CD₃OD) δ ppm 7.89 (d, J=1.37 Hz, 1H) 7.65 (d, J=1.37 Hz, 1H).

Acid Preparation 16 2-(3-methyl-1,2,4-oxadiazol-5-yl)isonicotinic acid

A mixture of acetonitrile (2 mol), hydroxylamine hydrochloride (2 mol)and sodium methoxide (2 mol) was stirred at room temperature for 3 days,then filtered and the filtrate concentrated below 20° C. to give(Z)-N′-hydroxyacetimidamide (150 g) as a white solid which was directlyused in the next step.

A mixture of methanol (800 mL), potassium hydroxide (44 g, 0.95 mol) anddimethyl pyridine-2,4-dicarboxylate (ChemPacific) (156 g, 0.79 mol) wasrefluxed for 0.5 hours and then evaporated in vacuo to afford4-(methoxycarbonyl)picolinic acid (144 g) as a yellow solid.

To 4-(methoxycarbonyl)picolinic acid (150 g, 1.62 mol) indichloromethane (500 mL) was added oxalyl chloride (400 mL) keeping thetemperature at 25-30° C. for 3 days. The reaction was evaporated invacuo to afford methyl 2-(chlorocarbonyl)isonicotinate as yellow oil.

To a solution of methyl 2-(chlorocarbonyl)isonicotinate indichloromethane (500 mL) was added (Z)-N′-hydroxyacetimidamide andtriethylamine, keeping the temperature at 25-30° C. for 1 day. Thereaction was concentrated in vacuo to afford (Z)-methyl2-((1-aminoethylideneaminooxy)carbonyl)isonicotinate as a yellow solid.

A solution of (Z)-methyl2-((1-aminoethylideneaminooxy)carbonyl)isonicotinate in toluene (1 L)was heated at reflux overnight. The obtained mixture was evaporated andpurified by silica-gel column chromatography to afford methyl2-(3-methyl-1,2,4-oxadiazol-5-yl)isonicotinate as a white solid.

A mixture of lithium hydroxide (15 g, 0.35 mol), ethanol (500 mL) andmethyl 2-(3-methyl-1,2,4-oxadiazol-5-yl)isonicotinate (52 g, 0.23 mol)were stirred at room temperature for 5 hours, then mixture wasconcentrated in vacuo. Water was added then extracted with ethylacetate. The water layer was brought to pH 1.5 with aqueous 1Nhydrochloric acid and extracted with ethyl acetate. The organic layerwas concentrated in vacuo to afford the title compound as a white solid(42 g). ¹H NMR (300 MHz, DMSO-d₆) δ ppm 14.08 (br s, 1H) 9.00-8.98 (m,1H) 8.50 (s, 1H) 8.09-8.07 (m, 1H), 2.46 (s, 3H).

Preparation I-1A-1a tert-butyl9-oxo-3-azaspiro[5.5]undec-7-ene-3-carboxylate

Methyl vinyl ketone (146 mL) was added to a solution of tert-butyl4-formylpiperidine-1-carboxylate (375 g) in tetrahydrofuran (18 L). Thereaction mixture was cooled to −5° C. and a solution of potassiumhydroxide in ethanol (3N, 0.243 L) was added dropwise over 10 minutes.The reaction mixture was allowed to warm to room temperature and stirredfor 16 hours. Cyclohexane (10 L) was added and the solution was washedwith saturated sodium chloride (3×10 L). The organic layer wasconcentrated to an oil. This oil was dissolved in 2 L of 80:20cyclohexane/ethyl acetate and filtered through Celite® to removeinsoluble material. The filtrate was purified via flash columnchromatography (70:30 hexane/ethyl acetate) to afford the product as anoil. The oil was triturated in hexanes to afford the desired product asa white solid (131 g, 28%).

Preparation I-1A-1b (E)-tert-butyl10-((dimethylamino)methylene)-9-oxo-3-azaspiro[5.5]undec-7-ene-3-carboxylate

Preparation I-1A-1a (101 g), and tris(dimethylaminomethane) (133 mL)were dissolved in toluene (800 mL) and heated to reflux for 17 hours.The reaction mixture was concentrated to a minimum stirring volume andethyl acetate (600 mL) was added. This mixture was heated to reflux andheptane (1.2 L) was added over 20 minutes. The hot solution was cooledto room temperature over 3 hours. The solids were filtered through acoarse glass frit and washed with heptane (300 mL). The resulting solidwas dried in a vacuum oven at 40° C. for 3 hours to afford the desiredproduct as a yellow solid (107 g). ¹H NMR (400 MHz, CDCl₃) δ ppm 7.48(s, 1H), 6.57 (d, J=9.97 Hz, 1H), 5.99 (d, J=10.16 Hz, 1H), 3.32-3.51(m, 4H), 3.06 (s, 6H), 2.72 (s, 2H), 1.57-1.66 (m, 2H), 1.41-1.53 (m,11H).

Preparation I-1A-1c tert-butyl1-isopropyl-1,4-dihydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-1A-1b (107 g) was taken up in toluene (700 mL) andisopropyl hydrazine (44.4 g) was added. The reaction was stirred atreflux for 4 hours. The reaction was cooled to room temperature andethyl acetate was added (500 mL). The reaction solution was washed withcitric acid (2×300 mL, 10% aqueous), and water (400 mL). The organiclayer concentrated in vacuo to afford I-1A-1c as a yellow solid (109 g).¹H NMR (400 MHz, CDCl₃) δ ppm 7.25 (s, 1H) 6.42 (dd, J=10.05, 0.49 Hz,1H) 5.84 (d, J=9.95 Hz, 1H) 4.42-4.52 (m, 1H) 3.36-3.53 (m, 4H) 2.62 (s,2H) 1.56-1.68 (m, 2H) 1.45-1.55 (m, 17H).

Preparation I-1A-1d tert-butyl1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

To a solution of Preparation I-1A-1c (109 g) in methanol (1 L) was addedN-bromo succinimide (61.4 g). The reaction was stirred for 1 hour. Thereaction was quenched with sodium thiosulfate (10 g in 300 mL water) andthen distilled to a final volume of 500 mL. The solution was cooled toambient temperature and 2-methyl tetrahydrofuran (1 L) and water (100mL) were added. The organic layer was removed and the aqueous layer wasextracted with 2-methyl tetrahydrofuran. The organic layers werecombined, washed with aqueous sodium hydroxide (1 N, 250 mL), water, andsaturated aqueous sodium chloride. The organic layer was dried oversodium sulfate, filtered and concentrated to an orange oil. The oil wasdissolved in tetrahydrofuran (500 mL) and potassium tert-butoxide (76.8g) in tetrahydrofuran (500 mL) was added. The solution was heated to 60°C. and stirred for 1 hour. Aqueous hydrochloric acid (1 N, 1 L) wasadded and the solution was stirred for 30 minutes. The phases wereseparated and the aqueous layer was extracted with ethyl acetate (700mL). The organic layers were combined, washed with water (400 mL) andsaturated aqueous sodium chloride (100 mL). The organic layer was driedover sodium sulfate, filtered and concentrated in vacuo to give aresidue. The residue was dried in a vacuum oven at 40° C. for 16 hoursto afford the title compound as an orange wax (117 g). ¹H NMR (400 MHz,CDCl₃) δ ppm 7.35 (s, 1H), 5.32-5.42 (m, 1H), 3.29-3.51 (m, 4H), 2.73(s, 2H), 2.51 (s, 2H), 1.47-1.57 (m, 4H), 1.42-1.46 (m, 15H); +ESI MS(M+H)=348.5.

Preparation I-1A-1e1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

Preparation I-1A-1d (23.5 g) was suspended in ethyl acetate (260 mL) andmethanol (70 mL). The reaction solution was cooled to <2° C. and acetylchloride (33.6 mL) was added dropwise over 20 min. The reaction wasallowed to slowly warm to room temperature and was stirred for 4 hours.The reaction solution was cooled to 0° C. and the precipitate wasfiltered. The precipitate was washed with ethyl acetate (200 mL) anddried in a vacuum oven (40° C., 10 mm Hg) for 16 hours to afford thetitle compound as a light yellow solid. ¹H NMR (400 MHz, CD₃OD) δ ppm7.43 (s, 1H), 5.32-5.42 (m, 1H), 3.15-3.25 (m, 4H), 2.89 (s, 2H), 2.64(s, 2H), 1.69-1.90 (m, 4H), 1.37-1.45 (m, 6H); +ESI MS (M+H)=248.4.

Alternate Preparation of1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-onePreparation (I-1A-1e)

tert-Butyl 9-oxo-3-azaspiro[5.5]undec-7-ene-3-carboxylate (250 g), andtris (dimethylaminomethane) (325 mL) were dissolved in toluene (1.9 L)and heated at reflux for 4 hours. The mixture was distilled andconcentrated to a minimum stirring volume (110° C.) and then toluene(1.9 L) was added. The reaction was redistilled to a minimum stirringvolume and cooled to room temperature. Toluene (1.8 L) and isopropylhydrazine hydrochloride (135 g) were added and the solution was heatedto reflux for 5 hours. The reaction was cooled to room temperature andwas then washed with citric acid (10% aqueous, 2×150 mL) and water (200mL), and then the organic layer was distilled to a minimum stirringvolume. Methanol (2 L) was added and distilled to a minimum stirringvolume. This was repeated with methanol (2 L). The solution wasredissolved in methanol (2.5 L) and N-bromosuccinimide (176 g) was addedin one portion. The solution was stirred at 23° C. for 2 hours. Aqueoussodium thiosulfate solution (5 wt %, 0.5 L) was added and the mixturewas stirred for 15 minutes. The reaction mixture was concentrated viadistillation (45° C., 210 mm Hg) to ˜0.5 L and then 2-methyltetrahydrofuran (2.5 L) was added. After stirring for 15 minutes theaqueous layer was discarded. The organic layer was concentrated to ˜0.2L and tetrahydrofuran (0.5 L) was added. To the mixture was added apotassium tert-butoxide solution in tetrahydrofuran (1.9 L, 1 Msolution). The solution was heated to 60° C. and stirred for 1 hour.After cooling to room temperature, aqueous hydrochloric acid (1 N, 2.2L) was added over 20 minutes. The mixture was stirred at roomtemperature for 20 minutes, and then the layers were allowed toseparate. The aqueous layer was removed and back extracted with ethylacetate (1.75 L). The combined organic layers were washed with water (1L) and the organic layer concentrated via distillation (4 L solventremoved). Ethyl acetate (1.8 L) was added and the solution wasconcentrated to a minimum stirring volume. Ethyl acetate (3 L) andmethanol (0.8 L) were added and the solution was cooled to 0° C. Acetylchloride (401 mL) was added dropwise over 20 minutes and the solutionwas stirred at 0° C. for 4 hours. The precipitate was collected byfiltration under nitrogen. The filtrate was washed with ethyl acetate(0.5 L) and dried in a vacuum oven at 40° C. to afford the I-1A-1e as anoff-white solid (241 g). ¹H NMR (400 MHz, CD₃OD) δ ppm 7.43 (s, 1H),5.32-5.42 (m, 1H), 3.15-3.25 (m, 4H), 2.89 (s, 2H), 2.64 (s, 2H),1.69-1.90 (m, 4H), 1.37-1.45 (m, 6H); +ESI (M+H)=248.4

Preparation I-IA-2 benzyl 9-oxo-3-azaspiro[5.5]undec-7-ene-3-carboxylate

To a benzene (700 mL) solution of benzyl4-formylpiperidine-1-carboxylate (90.0 g, 363.9 mmol) stirring in a 2 L3-neck flask fitted with a Dean-Stark trap was added p-toluenesulfonicacid (6.92 g, 36.4 mmol). The reaction was heated to 70° C.,3-buten-2-one (61.8 mL, 753 mmol) was added and mixture was heated atreflux for 24 hours collecting expelled water in the trap. The reactionwas cooled to room temperature and washed with 500 mL saturated aqueoussodium bicarbonate. The organic phase was dried over sodium sulfate,filtered and concentrated. The resultant dark brown oil was taken up in200 mL dichloromethane and filtered through a silica pad (600 mLsilica), eluting with 2 L heptane followed by 3 L 50% ethylacetate/heptane and then 3 L ethyl acetate, collecting by 1 L fractions.Fractions containing clean product were combined and concentrated toyield 68.1 g of the title compound as a thick brown oil. The fractionscontaining impure product were combined and concentrated and purified byflash chromatography (10-80% ethyl acetate/heptane, 340 g silica gel) toyield an additional 23.6 g of the title compound as a thick brown oil.Combined yield of 91.7 g, (94.1%) was realized. ¹H NMR (400 MHz, CDCl₃)δ ppm 7.27-7.43 (m, 5H), 6.79 (d, J=10.3 Hz, 1H), 5.95 (d, J=10.3 Hz,1H), 5.13 (s, 2H), 3.56-3.71 (m, 2H), 3.39-3.55 (m, 2H), 2.38-2.50 (m,2H), 1.96 (t, J=6.7 Hz, 2H), 1.70-1.52 (m, 4H).

Preparation I-1A-2a Benzyl10-((dimethylamino)methylene)-9-oxo-3-azaspiro[5.5]undec-7-ene-3-carboxylate

9-oxo-3-aza-spiro[5.5]undec-7-ene-3-carboxylic acid benzyl ester,Preparation I-IA-2 (15.2 g, 51 mmol) was dissolved in 180 mL toluene andtris(dimethylamino)methane (22.2 g, 27 mmol) was added. The reaction washeated to reflux for 5 hours and then allowed to cool to roomtemperature overnight. The reaction solution was concentrated in vacuoto provide the title compound (18.0 g, 100%): +APCI MS (M+H) 354.6; ¹HNMR (400 MHz, CDCl₃) δ ppm 7.49 (s, 1H), 7.28-7.40 (m, 5H), 6.59 (d,J=10.16 Hz, 1H), 6.01 (d, J=9.97 Hz, 1H), 5.13 (s, 2H), 3.52-3.66 (m,2H), 3.39-3.52 (m, 2H), 3.07 (s, 6H), 2.74 (s, 2H), 1.58-1.73 (m, 2H),1.41-1.58 (m, 2H).

Preparation I-1A-2b benzyl1-tert-butyl-1,4-dihydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-1A-2a (59.2 g, 167 mmol) was dissolved in 835 mL ofethanol. To the reaction solution was added acetic acid (20 mL, 345mmol) and tert-butylhydrazine hydrochloride (29.1 g, 234 mmol). Thereaction was heated to reflux for 1 hour. The reaction solution wascooled to room temperature and then concentrated in vacuo to give anorange oil which was purified by flash chromatography using 20-40% ethylacetate in heptane as eluent to afford the title compound as a paleyellow solid (50 g, 79%): +ESI MS (M+H) 380.5; ¹H NMR (400 MHz, CDCl₃) δppm 7.26-7.40 (m, 5H) 7.17 (s, 1H) 6.66 (d, J=9.95 Hz, 1H) 5.77 (d,J=10.15 Hz, 1H) 5.12 (s, 2H) 3.38-3.64 (m, 4H) 2.58 (s, 2H) 1.60 (s,12H) 1.50 (br. s., 1H).

Preparation I-1A-2c benzyl6-bromo-1-tert-butyl-7-hydroxy-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-1A-2b (50 g, 132 mmol) was dissolved in 1 L oftetrahydrofuran. To the reaction was added N-bromosuccinimide (24.6 g,138 mmol) and 250 mL of water. The reaction was stirred for 1 hour atroom temperature. The reaction was partitioned between ethyl acetate andwater. The phases were separated and the organic phase was washed anadditional 2 times with water and once with saturated aqueous sodiumchloride. The organic phase was dried over magnesium sulfate,concentrated in vacuo, and crystallized from ether to afford the titlecompound as a cream-colored solid (60.7 g, 97%): +ESI MS (M+H) 476.5; ¹HNMR (400 MHz, CDCl₃) δ ppm 7.28-7.36 (m, 5H), 7.27 (s, 1H), 5.23 (t,J=4.68 Hz, 1H), 5.12 (s, 2H), 4.24 (d, J=4.49 Hz, 1H), 3.87 (br. s.,2H), 3.12 (br. s., 2H), 2.79 (d, J=16.00 Hz, 2H), 2.59 (d, J=15.80 Hz,2H), 1.95 (br. s., 1H), 1.66 (s, 11H), 1.58 (br. s., 1H).

Preparation I-1A-2d benzyl6-bromo-1-tert-butyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-1A-2c (57.9 g, 122 mmol) was dissolved in 1 L acetone andthen cooled to 0° C. in an ice bath. To the solution was added 122 mL ofJones Reagent (Fillion, E. Tetrahedron Letters 2004, 46, 1091-1094). Theice bath was removed and the reaction was allowed to warm to roomtemperature where it was stirred for 45 minutes. Saturated, aqueoussodium bicarbonate was added until no further gas evolution was notedand pH reached 7. The resulting mixture was filtered through a pad ofCelite® rinsing with ethyl acetate. The filtrate layers were separatedand the aqueous layer was back extracted with ethyl acetate. The organicextracts were combined, washed twice with water, once with saturatedaqueous sodium chloride, dried over magnesium sulfate and concentratedin vacuo. The residue was crystallized from ethyl acetate/heptane toafford the title compound (50.4 g, 87%): +ESI MS (M+H) 474.5; ¹H NMR(400 MHz, CDCl₃) δ ppm 7.32 (d, J=9.38 Hz, 6H), 5.11 (s, 2H), 4.24 (s,1H), 3.58-3.84 (m, 2H), 3.16-3.41 (m, 2H), 2.67-2.91 (m, 2H), 1.80 (br.s., 1H), 1.61-1.76 (m, 11H), 1.52-1.61 (m, 1H).

Preparation I-1A-2e benzyl1-tert-butyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-1A-2d (50.4 g, 106 mmol) was dissolved in 600 mL oftetrahydrofuran, to this was added saturated aqueous ammonium chloride(600 mL) and zinc powder (20.8 g, 319 mmol). The reaction was stirredfor 30 minutes at room temperature. The reaction was filtered throughCelite®, the phases were separated and the organic phase was washed withwater and saturated aqueous sodium chloride, dried over magnesiumsulfate and concentrated in vacuo to give a foam. The foam wastriturated once in ethyl acetate/heptane and once in ether and filteredto afford the title compound as a white solid (40.4 g, 96%): +ESI MS(M+H) 396.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.24-7.38 (m, 6H), 5.11 (s,2H), 3.36-3.61 (m, 4H), 2.74 (s, 2H), 2.54 (s, 2H), 1.64 (s, 9H), 1.51(br. s., 4H).

Preparation I-1A-2f1-tert-butyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(I-1A-2f)

Preparation I-1A-2e (46.6 g, 118 mmol) was dissolved in 730 mL ethanoland the solution was added to 10% palladium on carbon (9.4 g). To thiswas added 1-methyl-1,4-cyclohexadiene (90 mL, 769 mmol). The reactionwas stirred at reflux for 2 hours. The reaction was cooled to roomtemperature and filtered through Celite®. The filtrate was concentratedin vacuo to give a gray solid. The solid was dissolved in 150 mL ethylacetate, to this was added 35 mL 4 M hydrochloric acid in dioxane. Aprecipitate formed and was collected by filtration to afford the titlecompound as a white solid (34 g, 97%): +ESI MS (M+H) 262.5; ¹H NMR (400MHz, CD₃OD) δ ppm 7.34 (s, 1H) 3.12-3.25 (m, 4H) 2.90 (s, 2H) 2.66 (s,2H) 1.67-1.85 (m, 4H) 1.62 (s, 9H).

Preparation I-2A-42a tert-butyl1-(2-ethoxy-2-oxoethyl)-1,4-dihydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Ethylhydrazinoacetate hydrochloride (0.92 g, 5.95 mmol) was added to asolution of Preparation I-1A-2a (1.25 g, 3.90 mmol) in ethanol (30 mL).Stir the mixture at reflux for 1 hour. An aliquot indicated the reactionwas complete by ¹HNMR. The reaction mixture was cooled to roomtemperature and concentrated under high vacuum to a brown oil. The oilwas triturated with diethyl ether (50 mL). The precipitate was filteredand the filtrate concentrated and dried under high vacuum to yield thetitle compound (1.50 g, 100%) as a to brown oil. +APCI MS (M+H) 376.2;¹H NMR (400 MHz, CDCl₃) δ ppm 1.21-1.26 (m, 3H), 1.43 (s, 9H), 1.45-1.52(m, 2H), 1.54-1.64 (m, 2H), 2.62 (s, 2H), 3.33-3.49 (m, 4H), 4.15-4.22(m, 2H), 4.82 (s, 2H), 5.87 (d, J=9.97 Hz, 1H), 6.26 (d, J=9.97 Hz, 1H),7.24 (s, 1H).

Preparation I-2A-42b diethyl2-(1″-(tert-butoxycarbonyl)spiro[indazole-5,4′-piperidine]-1(4H)-yl)malonate

Preparation I-2A-42a (1.45 g 3.86 mmol) in toluene (5 mL) was added to asuspension of sodium hydride (0.148 g, 60% dispersion in mineral oil) indiethyl carbonate (30 mL), dropwise at 80° C. over 30 minutes. Thereaction was stirred at reflux for 1.5 hours. ¹H NMR indicated thatstarting material was consumed and that the desired product had formed.The reaction mixture was cooled to room temperature. Methanol (1 mL) wasadded and the solution was stirred at room temperature for 5 minutes.Water (5 mL) was added. The solution was acidified to pH˜3 with 2 Naqueous, hydrochloric acid (3 mL) then was extracted withdichloromethane (3×15 mL). The combined organics were dried overmagnesium sulfate, filtered, concentrated, and dried under high vacuumto yield a brown gum (1.59 g, 92%). The crude material was trituratedwith 1:1 diethyl ether: heptanes (50 mL). The precipitate was filtered.The filtrate was concentrated and dried under high vacuum to yield thetitle compound (1.25 g, 72%). +APCI MS (M+H) 348.1; ¹H NMR (400 MHz,CDCl₃) δ ppm 1.13-1.32 (m, 6H), 1.40-1.46 (m, 9H), 1.46-1.54 (m, 2H),1.59 (d, J=13.68 Hz, 3H), 2.62 (s, 2H), 3.31-3.51 (m, 4H), 4.27 (q,J=7.23 Hz, 4H), 5.85 (d, J=9.97 Hz, 1H), 6.34 (d, J=9.97 Hz, 1H), 7.24(s, 1H).

Preparation I-2A-42c tert-butyl1-(1,3-dihydroxypropan-2-yl)-1,4-dihydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Tetrahydrofuran (40 mL) was added to lithium aluminum hydride (900 mg)in a 3-neck, 125 mL round bottom flask equipped with a nitrogen inletand thermometer. The solution was cooled to −2° C. Preparation I-2A-42b(1 g) in tetrahydrofuran (5 mL) was added dropwise over 5 minutes. Thetemperature was never greater than −0.2° C. during the addition. Thereaction was stirred at 0° C. for 3 hours then the reaction was thenquenched through the sequential addition of water (1.0 mL), 15% aqueoussodium hydroxide (1.0 mL), and water (3 mL). The internal temperaturewas never greater than 3.2° C. during the addition. The reaction wasthen allowed to warm to room temperature over 15 minutes. The reactionmixture was filtered through Celite® and washed with diethyl ether (3×20mL). The combined organics were washed with brine (5 mL) dried oversodium sulfate, filtered, concentrated, and dried under high vacuum toyield a pale yellow glass (548 mg, 67%). This material waschromatographed on 25 g of silica eluting with 2% to 8% methanol indichloromethane with 0.1% ammonium hydroxide over 30 minutes to yieldthe title compound (133 mg, 16%). +APCI MS (M+H) 364.2; ¹H NMR (400 MHz,CDCl₃) δ ppm 1.45 (s, 9H), 1.51 (br. s., 2H), 1.60 (br. s., 4H), 2.62(s, 2H), 3.32-3.53 (m, 4H), 4.05 (br. s., 4H), 4.26 (t, J=4.89 Hz, 1H),5.89 (s, 1H), 6.40 (d, J=9.77 Hz, 1H), 7.23-7.25 (m, 1H).

Preparation I-2A-42d tert-butyl1-(oxetan-3-yl)-1,4-dihydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

2.5 M n-butyl lithium in hexanes (0.33 ml, 165 uL) was added to asolution of Preparation I-2A-42c (150 mg, 0.41 mmol) in tetrahydrofuran(8 mL) at −6.2° C. The temperature was never greater than −3.7° C.during the addition. The solution was stirred at −8° C. for 30 minutes.A solution of p-toluenesulfonyl chloride (79 mg) in tetrahydrofuran (2mL) was added to the reaction mixture at −5° C. The temperature wasnever greater than −2° C. during the addition. The reaction was stirredat −5° C. for 1 hour then the reaction mixture was cooled to −6° C. and2.5 M n-butyl lithium in hexanes (0.33 mL, 165 uL) was added over 2minutes. The temperature was never greater than −3.5° C. during theaddition. The cooling bath was removed and the reaction was stirred atan internal temperature of 60° C. for 16 hours. The reaction mixture wascooled to room temperature and ethyl acetate (20 mL) was added. Thereaction solution was washed with water (35 mL) and the aqueous layerwas extracted with ethyl acetate (15 mL). The combined organics werewashed with brine (5 mL) dried over magnesium sulfate, filtered,concentrated, and dried under high vacuum to yield a yellow solid. Thesolid was purified by chromatography on 8 g silica eluting with 25% to75% ethyl acetate in heptanes over 36 minutes to yield the titlecompound (58 mg, 40%). +ESI MS (M+H); ¹H NMR (400 MHz, CDCl₃) δ ppm 1.45(s, 9H), 1.49 (d, J=3.71 Hz, 1H), 1.55 (s, 4H), 1.59 (br. s., 1H), 2.61(s, 2H), 3.32-3.50 (m, 4H), 5.00 (m, J=7.22, 7.22 Hz, 2H), 5.13 (t,J=6.44 Hz, 2H), 5.36-5.46 (m, 1H), 5.88 (d, J=9.95 Hz, 1H), 6.43 (d,J=9.95 Hz, 1H), 7.33 (s, 1H).

Preparation I-2A-42e tert-butyl6-bromo-7-methoxy-1-(oxetan-3-yl)-1,4,6,7-tetrahydrospiro[indazole-5,4″-piperidine]-1′-carboxylate

N-bromosuccinimide (30 mg, 0.17 mmol) was added to Preparation I-2A-42d(56 mg, 0.17 mmol) in methanol (1.0 mL) at room temperature. Thereaction was stirred at room temperature for 2 hours thenN-bromosuccinimide (4.5 mg) was added and the reaction mixture wasstirred at room temperature for 1 hour. The reaction mixture wasconcentrated under a stream of nitrogen to a residue. Ethyl acetate (15mL) was added and the reaction solution was washed with 10% citric acid(3 mL), 1N sodium hydroxide (3 mL), and brine (3 mL). The organic layerwas concentrated and dried under high vacuum to yield the title compound(74 mg, 100%) as a colorless solid. +APCI MS (M+H) 458.2; ¹H NMR (400MHz, CDCl₃) δ ppm 1.44 (s, 9H), 1.69 (br. s., 4H), 2.51 (d, J=15.83 Hz,1H), 2.67 (d, J=15.83 Hz, 1H), 3.06-3.31 (m, 3H), 3.54 (s, 3H),3.62-3.72 (m, 1H), 4.39 (s, 1H), 4.66 (s, 1H), 4.87-4.93 (m, 1H), 4.97(t, J=6.84 Hz, 1H), 4.99-5.04 (m, 1H), 5.30 (s, 1H), 5.34-5.40 (m, 1H),7.43 (s, 1H).

Preparation I-2A-42f tert-butyl1-(oxetan-3-yl)-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

1 M potassium tert-butoxide in tetrahydrofuran (0.320 mL) was added to asolution of Preparation I-2A-42e (72 mg, 0.16 mmol) in tetrahydrofuran(1.0 mL) at room temperature. The colorless solution turned yellow uponaddition. The solution was stirred at room temperature for 16 hours. 1 Naqueous, hydrogen chloride (0.475 mL, 3 eq.) was added and the solutionwas stirred at room temperature for 1 hour. The tetrahydrofuran wasconcentrated under a stream of nitrogen. The aqueous phase was extractedwith ethyl acetate (3×5 mL). The combined organics were washed withbrine (3 mL) then the organic layer was concentrated and dried underhigh vacuum to give the title compound as a pale yellow solid (54 mg,96%). −APCI MS (M−H) 360.2; ¹H NMR (400 MHz, CD₃OD) δ ppm 1.38-1.45 (m,9H), 1.46-1.56 (m, 4H), 2.57 (s, 2H), 2.82 (s, 2H), 3.33-3.53 (m, 4H),4.94-5.06 (m, 4H), 6.08-6.21 (m, 1H), 7.53 (s, 1H).

Preparation I-2A-42 g1-(oxetan-3-yl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

Trifluoroacetic acid (0.2 mL) was added to a solution of PreparationI-2A-42f (50 mg, 0.14 mmol) in dichloromethane (2 mL) at 0° C. Thecooling bath was removed and the reaction was stirred at roomtemperature for 1.5 hours. The reaction mixture was concentrated to aresidue under a stream of nitrogen and dried under high vacuum for 20minutes. The residue was triturated with diethyl ether (5 mL). Thesolvent was decanted and the resulting precipitate was dried under highvacuum to yield the title compound (52 mg, 100) as a pale yellow solid.+APCI MS (M+H) @ 262.2; ¹H NMR (400 MHz, CD₃OD) δ ppm 1.65-1.86 (m, 4H),2.63 (s, 2H), 2.89 (s, 2H), 3.14-3.27 (m, 4H), 5.02 (s, 4H), 6.07-6.21(m, 1H), 7.53-7.60 (m, 1H).

Preparation I-2A-75a Benzyl1-isopropyl-1,4-dihydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-1A-2a (6.38 g, 18 mmol) was dissolved in 90 mL of ethanol.To the reaction solution was added acetic acid (2.16 g, 36 mmol) and1-isopropylhydrazine hydrochloride (2.79 g, 25 mmol). The reaction washeated to reflux for 2 hours then the reaction solution was cooled toroom temperature and concentrated in vacuo to give an orange oil whichwas purified by flash chromatography using 12-100% ethyl acetate inheptane as eluent to afford the title compound as a yellow gum (6.58 g,69%): +ESI MS (M+H) 366.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.28-7.39 (m,5H), 7.25 (s, 1H), 6.42 (d, J=9.95 Hz, 1H), 5.84 (d, J=9.95 Hz, 1H),5.14 (s, 2H), 4.41-4.54 (m, 1H), 3.42-3.65 (m, 4H), 2.62 (s, 2H),1.58-1.70 (m, 2H), 1.50-1.58 (m, 2H), 1.49 (d, J=6.83 Hz, 6H).

Preparation I-2A-75b benzyl3,6-dibromo-1-isopropyl-7-methoxy-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-2A-75a (679 mg, 1.86 mmol) was dissolved in 15 mL methanoland treated with N-bromosuccinimide (728 mg, 4.09 mmol) and the reactionwas stirred at ambient temperature for 18 hours. The methanol wasremoved under reduced pressure. The resultant tan foam was taken up in50 mL ethyl acetate and washed with 0.5 M sodium hydroxide (2×50 mL) and20 mL saturated aqueous sodium thiosulfate. The organic phase was driedover sodium sulfate, filtered and concentrated. The resultant oil wasflash chromatographed (25 g silica, 10-80% ethyl acetate/heptanegradient) to yield 784 mg (76%) of the title compound as a white foam:+APCI-MS (M+H)=554.1, 556.2, 558.2: ¹H NMR (400 MHz, CDCl₃) δ ppm7.26-7.42 (m, 5H), 5.12 (s, 2H), 4.67 (d, J=1.76 Hz, 1H), 4.36 (s, 1H),4.27 (m, 1H), 3.79 (d, J=11.90 Hz, 1H), 3.59-3.73 (m, 1H), 3.53 (s, 3H),3.24-3.40 (m, 1H), 3.19 (ddd, J=13.61, 10.00, 3.12 Hz, 1H), 2.56 (d,J=16.19 Hz, 1H), 2.34 (d, J=16.19 Hz, 1H), 1.56-1.85 (m, 4H), 1.38-1.55(m, 6H).

Preparation I-2A-75c benzyl3-bromo-1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-2A-75b (784 mg, 1.4 mmol) was dissolved in 15 mLtetrahydrofuran. Potassium t-butoxide (2.82 mL, 2 eq, 1 Mtetrahydrofuran) was added and the reaction was stirred for 18 hours atambient temperature. To the reaction was added 25 mL 2 N hydrochloricacid. The mixture was stirred for 30 minutes at ambient temperature. Themixture was diluted with 25 mL water and extracted with ethyl acetate(2×50 mL). The organic extracts were combined and dried over sodiumsulfate, filtered and concentrated. The resultant oil was flashchromatographed (50 g silica, 8-66% ethyl acetate/heptane gradient) toyield 612 mg of the title compound as a white foam: +ESI MS (M+H)=462.5¹H NMR (400 MHz, CDCl₃) δ ppm 7.25-7.38 (m, 5H), 5.24-5.42 (m, 1H), 5.12(s, 2H), 3.49-3.66 (m, 2H), 3.46 (dd, J=7.41, 4.88 Hz, 2H), 2.63 (s,2H), 2.52 (s, 2H), 1.48-1.65 (m, 4H), 1.44 (d, J=6.63 Hz, 6H).

Preparation I-2A-75d tert-butyl3-bromo-1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-2A-75c (612 mg, 1.33 mmol) was dissolved in 10 mL 33%hydrobromic acid/acetic acid and the mixture was stirred for 60 minutesat ambient temperature. The solvent was evaporated and the red-orangeresidue taken up in 50 mL water and made basic with saturated aqueoussodium carbonate and extracted with ethyl acetate (2×50 mL). The organicphase was concentrated to 20 mL and treated with 20 mL saturated aqueoussodium bicarbonate and di-tert-butyl dicarbonate (348 mg). The biphasicmixture stirred for one hour at ambient temperature. The layers wereseparated and the organic phase dried over sodium sulfate, filtered andconcentrated under reduced pressure. The resultant oil was flashchromatographed (10-70% ethyl acetate/heptane, 10 g silica) to yield 364mg of the title compound. +ESI MS (M+H)=413.5; ¹H NMR (400 MHz, CDCl₃) δppm 5.24-5.43 (m, 1H) 3.41-3.56 (m, 2H) 3.28-3.41 (m, 2H) 2.63 (s, 2H)2.51 (s, 2H) 1.47-1.56 (m, 4H) 1.40-1.49 (m, 15H).

Preparation I-2A-75e tert-butyl1-isopropyl-3-methyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-2A-75d (440 mg, 1.03 mmol), palladium tetrakistriphenylphosphine (119 mg, 0.103 mmol), potassium carbonate (146 mg,1.03 mmol), and water (94 mg, 5.16 mmol) were combined indimethylformamide (2 mL) and degassed with nitrogen for 2 minutes. Thereaction vial was sealed and heated in a microwave reactor for 30minutes at 100° C. The vial was removed from the microwave reactor andthen heated to 100° C. in a conventional heating block for 4 days. Thereaction was concentrated in vacuo and then partitioned between water (5mL) and ethyl acetate (5 mL). The phases were separated and the organiclayer was concentrated and then chromatographed on a 40 g column elutingwith 20-40% ethyl acetate in heptane gradient to give 268 mg (72%) ofthe title compound. +ESI MS (M+H)=362.5; ¹H NMR (400 MHz, CDCl₃) δ ppm5.20-5.53 (m, 1H), 3.32-3.54 (m, 4H), 2.62 (s, 2H), 2.50 (s, 2H), 2.23(s, 3H), 1.53 (t, J=5.76 Hz, 4H), 1.46 (s, 9H), 1.44 (d, J=6.64 Hz, 6H).

Preparation I-2A-75f1-isopropyl-3-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

Preparation I-2A-75e (375 mg, 1.04 mmol) was dissolved in 3 mL diethylether and treated with 4 M hydrogen chloride in dioxane (1 mL). Thesolution was stirred for one hour and then concentrated in vacuo toprovide 300 mg of the title compound as a white foam: ¹H NMR (400 MHz,DMSO-d₆) δ ppm 5.10-5.35 (m, 1H), 4.34 (br. s., 4H), 2.70 (s, 2H), 2.56(s, 2H), 2.17 (s, 3H), 1.66 (br. s., 4H), 1.34 (d, J=6.64 Hz, 6H).

Preparation I-2A-76a tert-butyl3-cyano-1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

In a schlenk tube flushed with nitrogen was added Preparation I-2A-75d(250 mg, 0.59 mmol), tris(dibenzylideneacetone)dipalladium(0)-chloroformadduct (23.8 mg, 0.02 mmol), zinc dust (9.6 mg, 0.15 mmol), zinc cyanide(75.7 mg, 0.65 mmol) and 1,1′-bis(diphenylphosphino)ferrocene (26.1 mg,0.05 mmol). Anhydrous dimethylacetamide (3.5 mL) was added and the flaskwas flushed with nitrogen, then capped with a Teflon® screw top. Thereaction was stirred at 120° C. for 16 hours. The reaction was cooledand then filtered through a pad of Celite® washing with ethyl acetate.The filtrate was washed with water and the aqueous phase was backextracted with ethyl acetate. The combined organic phases were washedwith saturated aqueous sodium chloride, dried over magnesium sulfate andconcentrated in vacuo. The residue was purified by flash chromatographyusing 5-30% ethyl acetate in heptane gradient to give 204 mg of thetitle compound as a solid (93%): +ESI MS (M-Boc+H) 273.5; ¹H NMR (400MHz, CD₃OD) δ ppm 5.44 (m, 1H), 3.44 (m, 4H), 2.89 (s, 2H), 2.64 (s,2H), 1.53 (m, 4H), 1.46-1.43 (m, 15H).

Preparation I-8a-1 b1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-3-carbonitrile

Preparation I-2A-76a (70 mg, 0.19 mmol) was dissolved in dichloromethane(3 mL) and trifluoroacetic acid (0.2 mL) and stirred at ambienttemperature for 90 minutes. The solvent was concentrated in vacuo andthe residue was co-distilled with toluene followed by ethyl acetate togive 149 mg (100%) of the title compound as a yellow solid: +ESI MS(M+H) 273.5.

Preparation I-4A-1a Benzyl6-bromo-7-hydroxy-1-isopropyl-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-2A-75a (4.20 g, 11 mmol) was dissolved in 130 mL oftetrahydrofuran. To the reaction was added N-bromosuccinimide (2.49 g,14 mmol) and 30 mL of water. The reaction was stirred for 1 hour at roomtemperature. The reaction was partitioned between ethyl acetate andsaturated, aqueous sodium chloride. The organic phase was separated thenwashed an additional time with saturated aqueous sodium chloride. Theorganic phase was dried over sodium sulfate and concentrated in vacuo toafford the title compound as an off-white foam (5.31 g, 100%): +ESI MS(M+H) 463.8.

Preparation I-4A-1b Benzyl6-bromo-1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-4A-1a (5.30 g, 11 mmol) was dissolved in 53 mL acetone andthen cooled to 0° C. in an ice bath. To the solution was added 83 mL ofJones Reagent (Fillion, E. Tetrahedron Letters 2004, 46, 1091-1094). Theice bath was removed and the reaction was allowed to warm to roomtemperature where it was stirred for 45 minutes. The reaction was cooledto 0° C. in an ice bath and then saturated, aqueous sodium bicarbonatewas added until no further gas evolution was noted. The resultingmixture was filtered through a pad of Celite® rinsing with ethylacetate. The filtrate layers were separated and the aqueous layer wasback extracted with ethyl acetate. The organic extracts were combined,washed twice with water, once with saturated, aqueous sodium chloride,dried over sodium sulfate and concentrated in vacuo to afford the titlecompound (5.27 g, 100%): +ESI MS (M+H) 460.4.

Preparation I-4A-1c Benzyl6-bromo-1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-4A-1b (5.63 g, 12 mmol) was dissolved in 55 mL of aceticacid, to this was added zinc powder (2.40 g, 37 mmol). The reaction wasstirred for 35 minutes at room temperature. The reaction wasconcentrated in vacuo and then partitioned between saturated, aqueoussodium bicarbonate and ethyl acetate. The phases were separated and theaqueous phase was extracted with ethyl acetate. The organic extractswere combined, washed with water, saturated, aqueous sodium chloride,dried over sodium sulfate and concentrated in vacuo to give an oil. Theoil was purified by flash chromatography using 12-100% ethyl acetate inheptane as eluent to afford the title compound as an oil (2.25 g, 48%):+ESI MS (M+H) 382.4; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.28-7.40 (m, 6H),5.32-5.45 (m, 1H), 5.13 (s, 2H), 3.41-3.61 (m, 4H), 2.76 (s, 2H), 2.54(s, 2H), 1.50-1.62 (m, 4H), 1.47 (d, J=6.63 Hz, 6H).

Preparation I-4A-1d Benzyl1-isopropyl-6-methyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-4A-1c (397 mg, 1.04 mmol) in tetrahydrofuran (8 mL) wascooled to −70° C. To this was added lithium bis(trimethylsilyl)amide(1.56 mL, 1.56 mmol) as a 1.0 M solution in tetrahydrofuran over a tenminute period. The resulting yellow solution was stirred for thirtyminutes at −70° C. 1,3-Dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone(1.6 mL) was added to the reaction, stirring was continued at −70° C.for ten minutes. To the reaction was added iodomethane (746 mg, 5.2mmol). The reaction was allowed to warm to room temperature where it wasstirred for 18 hours. To the reaction was added saturated, aqueoussodium bicarbonate (2 mL), the mixture was then partitioned betweenwater (20 mL) and ethyl acetate (150 mL). The layers were separated andthe aqueous layer was extracted with ethyl acetate (150 mL). The organiclayers were combined, dried over magnesium sulfate, filtered and thenconcentrated to give a clear oil. The oil was purified by silica gelchromatography using 10-40% ethyl acetate in heptane as eluent to affordthe title compound as a white solid (351 mg, 85%): +ESI MS (M+H) 396.2;¹H NMR (400 MHz, DMSO-d₆) δ ppm 7.44 (s, 1H), 7.35 (s, 5H), 5.17-5.34(m, 1H), 5.06 (s, 2H), 3.52-3.72 (m, 4H), 2.79 (s, 2H), 2.42-2.48 (m,1H), 1.38-1.49 (m, 4H), 1.35 (t, J=6.74 Hz, 6H), 1.04 (d, J=7.04 Hz,3H).

Preparation I-4A-1e1-isopropyl-6-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

The title compound was prepared from Preparation I-4A-1d in an analogousfashion to Preparation I-1A-2f.

Preparation I-6A-1a Benzyl1-isopropyl-6,6-dimethyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

A solution of Preparation I-4A-1d (84 mg, 0.21 mmol) in 1 mLtetrahydrofuran was cooled to −70° C. and then treated with lithiumbis(trimethylsilyl)amide (0.318 mL, 0.318 mmol) as a 1.0 M solution intetrahydrofuran over ten minutes. Then1,3-Dimethyl-3,4,5,6-tetrahydro-2(1H)-pyrimidinone (0.2 mL) was added tothe reaction. Stirring continued for ten minutes at −70° C., theniodomethane (152 mg, 1.06 mmol) was added to the reaction. The mixturewas allowed to warm to room temperature where it was held for fourhours. To the reaction was added saturated, aqueous ammonium chloride (1mL), the mixture was then partitioned between water (2 mL) and ethylacetate (10 mL). The layers were separated and the aqueous layer wasextracted with ethyl acetate (5 mL). The organic layers were combined,dried over magnesium sulfate, filtered and then concentrated to give aclear, yellow oil. The oil was purified by silica gel chromatographyusing 10-40% ethyl acetate in heptane as eluent to afford the titlecompound as a clear oil (58 mg, 67%): +ESI MS (M+H) 410.3; ¹H NMR (400MHz, CDCl₃) δ ppm 7.28-7.44 (m, 5H), 7.27 (s, 1H), 5.40 (m, 1H), 5.13(s, 2H), 3.85-4.24 (m, 2H), 2.86-3.11 (m, 2H), 1.58-1.79 (m, 2H), 1.56(s, 2H), 1.46 (d, J=6.64 Hz, 6H), 1.19-1.40 (m, 2H), 1.15 (s, 6H).

Preparation I-6A-1b1-isopropyl-6,6-dimethyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

Preparation I-6A-1b was prepared from Preparation I-6A-1a in ananalogous fashion to Preparation I-1A-2f.

Preparation I-13A-1a tert-butyl1-tert-butyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

The hydrochloride salt of Preparation I-1A-2f (1040 mg, 3.492 mmol),di-tert-butyl dicarbonate (800 mg, 3.67 mmol) and triethylamine (730 mg,7.2 mmol) were combined in dichloromethane (30 mL). The reactionsolution was stirred at ambient temperature for 16 hours. To thereaction was added dichloromethane (20 mL). The reaction solution waswashed with 1N aqueous hydrochloric acid (5 mL), water (5 mL), andsaturated, aqueous sodium chloride (5 mL). The organic phase was driedover magnesium sulfate and concentrated to give I-13A-1a (1262 mg,100%): −APCI MS (M−H) 360.3; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.30 (s, 1H),3.29-3.56 (m, 4H), 2.77 (s, 2H), 2.56 (s, 2H), 1.67 (s, 9H), 1.48-1.56(m, 4H), 1.46 (s, 9H).

Preparation I-13A-1b tert-butyl3-bromo-1-tert-butyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-1′-carboxylate

Preparation I-13A-1a (1090 mg, 3.015 mmol) and sodium acetate (1050 mg,12.80 mmol) were combined in ethanol (40 mL) and water (10 mL). To thissolution was added bromine (1870 mg, 11.7 mmol). The reaction wasstirred at room temperature for 4 hours. To the reaction was addedethanol (40 mL). The reaction was stirred for 16 more hours. Thereaction solution was poured in water (20 mL) and extracted twice withethyl acetate (75 mL each). The combined organic extracts were washedtwice with aqueous, saturated sodium thiosulfate (25 mL each) andsaturated, aqueous sodium chloride (25 mL). The organic phase was driedover magnesium sulfate and concentrated to a final volume of 20 mL togive a precipitate. The mixture was filtered and the solids collected togive the title compound as a solid (679 mg, 51%): +APCI MS (M+H-Boc)342.1; ¹H NMR (400 MHz, CDCl₃) δ ppm 3.28-3.60 (m, 4H), 2.66 (s, 2H),2.56 (s, 2H), 1.65 (s, 9H), 1.48-1.55 (m, 4H), 1.46 (s, 9H).

Preparation I-13A-1c3-bromo-1-tert-butyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

Preparation I-13A-1b (670 mg, 1.52 mmol) and 4 M hydrogen chloride indioxane (8 mL) were combined and stirred for 2.5 hours. To the reactionwas added diethyl ether (20 mL). A precipitate formed that was filteredand the solids collected to give 1-13A-1c (573 mg, 97%): +APCI MS (M+H)342.1; ¹H NMR (400 MHz, CD₃OD) δ ppm 3.24 (t, J=5.96 Hz, 4H), 2.80 (s,2H), 2.74 (s, 2H), 1.71-1.92 (m, 4H), 1.65 (s, 9H).

Example 1 Preparation of1-isopropyl-1′-(4-chloro-3-methylbenzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(1A-1)

4-chloro-3-methylbenzoic acid (253 mg, 1.48 mmol) was suspended inthionyl chloride (3000 mg, 30 mmol) and heated to reflux for 30 minutes.The solution was concentrated in vacuo in the presence ofdichloromethane to give a residue. The residue was dissolved in 1 mLdichloromethane and added to a solution of diisopropyl ethylamine (890mg, 6.9 mmol) and Preparation I-1A-1e (204 mg, 0.83 mmol) in 8 mLdichloromethane. The reaction was stirred for 10 minutes. The reactionwas partitioned between dichloromethane and saturated, aqueous sodiumbicarbonate. The organic phase as separated and then washed withsaturated, aqueous sodium chloride. The organic layer was dried oversodium sulfate and concentrated in vacuo to give an oil. The oil waspurified by flash chromatography using 15-100% ethyl acetate in heptaneas eluent to afford the title compound as a white foam (153 mg, 46%):+ESI MS (M+H) 400.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.32-7.40 (m, 2H),7.28 (s, 1H), 7.11-7.17 (m, 1H), 5.31-5.45 (m, 1H), 3.74 (br. s., 2H),3.42 (br. s., 2H), 2.80 (s, 2H), 2.59 (s, 2H), 2.39 (s, 3H), 1.49-1.84(m, 4H), 1.46 (d, J=6.63 Hz, 6H).

The compounds listed in Table 1 below were prepared using proceduresanalogous to those described above for the synthesis of Example 1,Compound 1A-1 using the appropriate starting materials which areavailable commercially, prepared using preparations well-known to thoseskilled in the art, or prepared in a manner analogous to routesdescribed above for other intermediates.

TABLE 1

Ex. R¹ Analytical Data 1A-2 C(CH₃)₃ +ESI MS (M + H) 414.1; ¹H NMR (400MHz, CDCl₃) δ ppm 7.35 (d, J = 8.19 Hz, 1 H) 7.31 (s, 1 H) 7.28 (d, J =2.15 Hz, 1 H) 7.14 (dd, J = 7.80, 1.95 Hz, 1 H) 3.19- 4.01 (m, 4 H) 2.82(s, 2 H) 2.61 (s, 2 H) 2.39 (s, 3 H) 1.66 (s, 9 H) 1.58 (s, 4 H) 1A-3cyclobutyl +ESI MS (M + H) 412.4; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.39 (s,1 H) 7.31- 7.37 (m, 1 H) 7.25 (s, 1 H) 7.12 (dd, J = 8.01, 1.76 Hz, 1 H)5.58 (quin, J = 8.35 Hz, 1 H) 3.72 (br. s., 2 H) 3.38 (br. s., 2 H) 2.77(s, 2 H) 2.56-2.68 (m, 2 H) 2.55 (s, 2 H) 2.38-2.46 (m, 2 H) 2.37 (s, 3H) 1.74 - 1.93 (m, 2 H) 1.40- 1.72(m, 4 H) 1A-4 benzyl +ESI MS (M + H)448.4; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.39 (s, 1 H) 7.31- 7.36 (m, 2 H)7.21-7.29 (m, 5 H) 7.09- 7.15 (m, 1 H) 5.65 (s, 2 H) 3.52-3.89 (m, 2 H)3.15-3.52 (m, 2 H) 2.78 (s, 2 H) 2.55 (s, 2 H) 2.37 (s, 3 H) 1.41-1.75(m, 4 H) 1A-5

+ESI MS (M + H) 428.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.38 (s, 1 H) 7.34(d, J8.00 Hz, 1 H) 7.25-7.27 (m, 1 H) 7.10-7.14 (m, 1 H) 5.73-5.82 (m, 1H) 4.04-4.19 (m, 2 H) 3.89-3.99 (m, 2 H) 3.72 (br. s., 2 H) 3.39 (br.s., 2 H) 2.79 (s, 2 H) 2.58 (s, 2 H) 2.38-2.47 (m, 2 H) 2.36 (s, 3 H)1.58 (s, 4 H)

Example 2 Preparation of1-Isopropyl-1′-(1H-pyrazolo[3,4-b]pyridine-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(2A-1)

The hydrochloride salt of Preparation I-1A-1e (80 mg, 0.28 mmol),1H-pyrazolo[3,4-b]pyridine-5-carboxylic acid (46 mg, 0.28 mmol),O-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (107 mg, 0.28 mmol) and triethylamine (115 mg, 1.13mmol) were combined in 3 mL of dimethylformamide and stirred at roomtemperature for 16 hours. The reaction was partitioned between 10 mLethyl acetate and 10 mL saturated aqueous sodium bicarbonate. Theorganic phase was separated and then concentrated to give an oil. Theoil was purified by flash chromatography using 50-100% ethyl acetate inheptane as eluent to afford the title compound as a solid (48 mg, 44%):+APCI MS (M+H) 393.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 11.08 (br. s., 1H),8.67 (d, J=1.95 Hz, 1H), 8.22 (d, J=1.76 Hz, 1H), 8.16 (s, 1H), 7.40 (s,1H), 5.32-5.46 (m, 1H), 3.24-4.13 (m, 4H), 2.84 (s, 2H), 2.63 (s, 2H),1.66 (br. s., 4H), 1.47 (d, J=6.63 Hz, 6H).

The compounds listed in Table 2 below were prepared using proceduresanalogous to those described above for the synthesis of Compound 2A-1using the appropriate starting materials which are availablecommercially, prepared using preparations well-known to those skilled inthe art, or prepared in a manner analogous to routes described above forother intermediates. The compounds listed below were isolated initiallyas the free base and may be converted to their correspondinghydrochloride salt for testing.

TABLE 2

Ex. R¹ R² C(O)R⁴ Analytical Data 2A-2

H

+APCI MS (M + H) 442.2; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 7.42-7.49 (m, 2H) 7.37 (d, J = 1.76 Hz, 1 H) 7.18-7.25 (m, 1 H) 4.51 (dd, J = 13.27,7.22 Hz, 1 H) 4.32 (dd, J = 13.36, 5.37 Hz, 1 H) 4.07-4.17 (m, 1 H)3.62- 3.75 (m, 2 H) 3.53-3.62 (m, 2 H) 3.24-3.33 (m, 2 H) 2.80 (s, 2 H)2.60 (s, 2 H) 2.34 (s, 3 H) 1.68-1.91 (m, 3 H) 1.37-1.66 (m, 5 H) 2A-3CH(CH₃)CH₂OCH₃ H

+ESI MS (M + H) 430.1; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 7.41-7.51 (m, 2H) 7.37 (d, J = 2.15 Hz, 1 H) 7.21 (dd, J = 7.90, 1.85 Hz, 1 H) 4.51(dd, J = 13.27, 7.22 Hz, 1 H) 4.32 (dd, J = 13.46, 5.27 Hz, 1 H) 4.09-4.17 (m, 1 H) 3.62-3.74 (m, 2 H) 3.44-3.61 (m, 2 H) 3.31 (s, 3 H) 2.80(s, 2 H) 2.59 (s, 2 H) 2.34 (s, 3 H) 1.67-1.90 (m, 2 H) 1.37-1.65 (m, 5H) 2A-4 CH₃ H

+ESI (MS) 372.1; ¹H NMR (400 MHz, CD₃OD) δ ppm 7.43 (d, J = 8.19 Hz, 1H) 7.39 (s, 1 H) 7.34 (d, J = 1.56 Hz, 1 H) 7.22 (d, J = 1.56 Hz, 1 H)4.07 (s, 3 H) 3.56-3.95 (m, 2 H) 3.35-3.55 (m, 2 H) 2.87 (s, 2 H) 2.63(d, J = 1.95 Hz, 2 H) 2.40 (s, 3 H) 1.41-1.83 (m, 4 H) 2A-5 CH₂CH(CH₃)₂H

+ESI (MS) 414.4; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.30-7.43 (m, 2 H) 7.28(s, 1 H) 7.14 (dd, J = 8.11, 2.05 Hz, 1 H) 4.31 (d, J = 7.43 Hz, 2 H)3.16-3.99 (m, 4 H) 2.81 (s, 2 H) 2.58 (s, 2 H) 2.39 (s, 3 H) 2.07-2.28(m, 1 H) 1.66 (s, 4 H) 0.87 (d, J = 6.84 Hz, 6 H) 2A-6

H

+ESI (MS) 462.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.40 (s, 1 H) 7.14-7.36(m, 7 H) 7.10 (dd, J = 8.11, 1.86 Hz, 1 H) 6.42 (q, J = 7.04 Hz, 1 H)3.10-3.97 (m, 4 H) 2.64-2.88 (m, 2 H) 2.42- 2.63 (m, 2 H) 2.36 (s, 3 H)1.86 (d, J = 7.04 Hz, 3 H) 1.61 (s, 4 H) 2A-7 CH₂CH₃ H

+ESI (MS) 386.5 2A-8 CH₂CH₂OCH₃ H

+APCI MS (M + H) 416.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.40 (s, 1 H) 7.36(d, J = 8.19 Hz, 1 H) 7.28 (d, J = 1.76 Hz, 1 H) 7.14 (dd, J = 8.00,1.76 Hz, 1 H) 4.69 (t, J = 5.56 Hz, 2 H) 3.75 (t, J = 5.56 Hz, 4 H)3.33-3.54 (m, 2 H) 3.31 (s, 3 H) 2.81 (s, 2 H) 2.59 (s, 2 H) 2.39 (s, 3H) 1.62 (s, 4 H) 2A-9 (CH₂)₂CH₃ H

+APCI MS (M + H) 400.2; ¹H NMR (400 MHz, CD₃OD) δ ppm 7.39-7.49 (m, 2 H)7.33-7.36 (m, 1 H) 7.19-7.25 (m, 1 H) 4.43 (t, J = 7.02 Hz, 2 H) 3.76-3.95 (m, 1 H) 3.60-3.76 (m, 1 H) 3.36-3.54 (m, 2 H) 2.89 (s, 2 H) 2.64(d, J = 1.95 Hz, 2 H) 2.40 (s, 3 H) 1.79 (sxt, J = 7.26 Hz, 2 H)1.47-1.73 (m, 4 H) 0.85 (t, J = 7.41 Hz, 3 H 2A-10 CH(CH₃)₂ H

+ESI MS (M + H) 406.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.98 (d, J = 0.78Hz, 1 H) 7.71 (d, J = 7.82 Hz, 1 H) 7.48 (d, J = 0.78 Hz, 1 H) 7.36 (s,1 H) 7.07-7.13 (m, 1 H) 5.30-5.39 (m, 1 H) 4.07 (s, 3 H) 3.80 (br. s., 2H) 3.42 (br. s., 2 H) 2.79 (s, 2 H) 2.58 (s, 2 H) 1.69 (br. s., 2 H)1.54 (br. s., 2 H) 1.43 (d, J = 6.64 Hz, 6 H) 2A-11 CH(CH₃)₂ H

+ESI MS (M + H) 406.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 8.00 (s, 1 H) 7.79(s, 1 H) 7.32-7.47 (m, 3 H) 5.30-5.41 (m, 1 H) 4.07 (s, 3 H) 3.61 (br.s., 4 H) 2.79 (s, 2 H) 2.58 (s, 2 H) 1.61 (br. s., 2 H) 1.53 (br. s., 2H) 1.43 (d, J = 6.84 Hz, 6 H) 2A-12 CH(CH₃)₂ H

+ESI MS (M + H) 392.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.99 (s, 1 H) 7.69(s, 1 H) 7.58 (d, J = 7.61 Hz, 1 H) 7.39 (s, 1 H) 7.29 (dd, J = 8.19,1.37 Hz, 1 H) 5.38 (quin, J = 6.63 Hz, 1 H) 3.35- 3.97 (m, 4 H) 2.82 (d,J = 1.95 Hz, 2 H) 2.61 (s, 2 H) 1.50-1.82 (m, 4 H) 1.46 (d, J = 6.63 Hz,6 H) 2A-13 CH(CH₃)₂ H

+ESI (MS) 419.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 9.59 (br. s., 1 H) 8.41(d, J = 1.76 Hz, 1 H) 7.76 (dd, J = 8.19, 1.76 Hz, 1 H) 7.61 (d, J =8.00 Hz, 1 H) 7.39 (s, 1 H) 7.15 (dd, J = 7.12, 5.76 Hz, 1 H) 6.56 (d, J= 7.41 Hz, 1 H) 5.38 (qd, J=6.63, 6.44 Hz, 1 H) 3.64- 4.02 (m, 2 H)3.34-3.64 (m, 2 H) 2.82 (s, 2 H) 2.61 (s, 2 H) 1.52-1.91 (m, 4 H) 1.46(d, J = 6.44 Hz, 6 H) 2A-14 CH(CH₃)₂ H

+ESI (MS) 419.4; ¹H NMR (400 MHz, CDCl₃) δ ppm 8.44 (d, J = 8.19 Hz, 1H) 7.60 (d, J = 1.37 Hz, 1 H) 7.47 (dd, J = 8.19, 1.56 Hz, 1 H) 7.39 (s,1 H) 7.18 (ddd, J = 7.07, 1.90, 1.17 Hz, 1 H) 6.58 (d, J = 7.22 Hz, 1 H)5.38 (quin, J = 6.63 Hz, 1 H) 3.66-3.96 (m, 2 H) 3.40 (br. s., 2 H) 2.82(s, 2 H) 2.61 (s, 2 H) 1.72 (br. s., 2 H) 1.51-1.61 (m, 2 H) 1.45 (s, 6H) 2A-15 CH(CH₃)₂ H

+ESI (MS) 383.5; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.05 (s, 1 H) 7.42 (s, 1H) 5.24-5.58 (m, 1 H) 3.37-3.96 (m, 4 H) 2.87 (s, 2 H) 2.63 (s, 2 H)2.43 (s, 3 H) 1.63 (br. s., 4 H) 1.42 (d, J = 6.64 Hz, 6 H) 2A-16CH(CH₃)₂ H

+ESI MS (M + H) 426.4; ¹H NMR (400 MHz, CDCl₃) δ ppm 10.57 (br. s., 1 H)8.14 (s, 1 H) 7.72 (d, J = 1.17 Hz, 1 H) 7.45 (d, J = 1.17 Hz, 1 H) 7.37(s, 1 H) 5.30-5.42 (m, 1 H) 3.59 (br. s., 4 H) 2.80 (s, 2 H) 2.59 (s, 2H) 1.62 (br. s., 4 H) 1.44 (d, J = 6.63 Hz, 6 H) 2A-17 CH(CH₃)₂ H

+ESI MS (M + H) 382.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 8.20 (dd, J = 2.34,0.59 Hz, 1 H), 7.56 (dd, J = 8.58, 2.34 Hz, 1 H) 7.38 (s, 1 H), 6.39(dd, J = 8.68, 0.68 Hz, 1H) 5.31-5.47 (m, 1 H), 4.83 (q, J = 5.14 Hz, 1H), 3.51- 3.69 (m, 4 H), 2.96 (d, J = 5.27 Hz, 3 H), 2.80 (s, 2 H) 2.59(s, 2 H), 1.53-1.75 (m, 4 H) 2A-18 CH(CH₃)₂ H

+ESI MS (M + H) 410.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.94 (s, 1 H) 7.35(s, 1 H) 6.30 (s, 1 H) 5.30-5.40 (m, 1 H) 3.26- 3.83 (br. m, 4 H) 3.06(s, 6 H) 2.77 (s, 2 H) 2.55 (s, 2 H) 2.23 (s, 3 H) 1.56 (br. s., 4 H)1.37- 1.47 (m, 6 H) 2A-19 CH(CH₃)₂ H

+APCI MS (M + H) 392.2; ¹H NMR (400 MHz, DMSO-d₆), δ ppm 11.76 (br. s.,1 H), 8.43 (d, J = 1.76 Hz, 1 H) 7.95 (s, 1 H) 7.86 (t, J = 2.93 Hz, 'IH) 7.46 (s, 1 H) 6.65 (ddd, J = 2.98, 1.90, 0.78 Hz, 1 H) 5.27 (quin, J= 6.58 Hz, 1 H) 3.26-3.86 (m, 4 H) 2.82 (s, 2 H) 2.63 (s, 2 H) 1.41-1.66 (m, 4 H) 2A-20 CH(CH₃)₂ H

+ESI MS (M + H) 392.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 12.05 (s, 1 H)8.30 (dd, J = 4.69, 1.56 Hz, 1 H) 8.00 (dd, J = 7.82, 1.56 Hz, 1 H) 7.47(s, 1 H) 7.03- 7.16 (m, 1 H) 6.71 (d, J = 2.15 Hz, 1 H) 5.28 (quin, J =6.64 Hz, 1 H) 3.57-3.82 (m, 4 H) 2.83 (s, 2 H) 2.64 (s, 2 H) 1.52-1.61(m, 4 H) 1.37 (d, J = 6.64 Hz, 6 H) 2A-21 C(CH₃)₃ H

+ESI MS (M + H) 420.5; ¹H NMR (400 MHz, CD₃OD) δ ppm 7.56 (d, J = 0.78Hz, 1 H) 7.54 (d, J = 8.21 Hz, 1 H) 7.34 (s, 1 H) 7.27 (dd, J = 8.21,1.56 Hz, 1 H) 3.88 (br. s., 1 H) 3.71 (br. s., 1 H) 3.45-3.58 (m, 2 H)2.90 (s, 2 H) 2.66 (s, 2 H) 2.59 (s, 3 H) 1.65-1.81 (m, 2 H) 1.63 (s, 9H) 1.51-1.60 (m, 2 H) 2A-22 C(CH₃)₃ H

+ESI (MS) 433.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 11.53 (br. s., 1 H) 8.39(d, J = 8.21 Hz, 1 H) 7.55 (d, J = 1.17 Hz, 1 H) 7.34- 7.48 (m, 1 H)7.27 (s, 1 H) 7.11- 7.22 (m, 1 H) 6.52 (d, J = 7.23 Hz, 1 H) 3.58-3.98(m, 2 H) 3.19-3.53 (m, 2 H) 2.79 (s, 2 H) 2.59 (s, 2 H) 1.63-1.79 (m, 2H) 1.61 (s, 9 H) 1.39-1.55 (m, 2 H) 2A-23 C(CH₃)₃ H

+ESI (MS) 406.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 10.95 (br. s., 1 H) 8.44(d, J = 1.95 Hz, 1 H) 8.04 (d, J = 1.95 Hz, 1 H) 7.38- 7.50 (m, 1 H)7.31 (s, 1 H) 6.55 (dd, J = 3.52, 1.95 Hz, 1 H) 3.22- 4.13 (m, 4 H) 2.83(s, 2 H) 2.63 (s, 2 H) 1.65 (s, 13 H) 2A-24 C(CH₃)₃ H

+ESI (MS) 406.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 10.05 (br. s., 1 H) 8.49(d, J = 1.76 Hz, 1 H) 7.68 (d, J = 0.78 Hz, 1 H) 7.49 (t, J = 3.02 Hz, 1H) 7.32 (s, 1 H) 6.70 (ddd, J = 3.07, 2.00, 0.78 Hz, 1 H) 3.29-4.03 (m,4 H) 2.83 (s, 2 H) 2.62 (s, 2 H) 1.66 (s, 13 H) 2A-25 C(CH₃)₃ H

+APCI (M + H) 440.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 8.16 (s, 1 H) 7.74(d, J = 1.17 Hz, 1 H) 7.48 (d, J = 1.17 Hz, 1 H) 7.32 (s, 1 H) 3.64 (br.s., 4 H) 2.84 (s, 2 H) 2.64 (s, 2 H) 1.67 (br, s., 13 H) 2A-26 CH(CH₃)₂H

+APCI (M + H) 392.2; ¹H NMR (400 MHz, CD₃OD) δ ppm 1.40 (d, J = 6.63 Hz,6 H), 1.53-1.74 (m, 4 H), 2.64 (s, 2 H), 2.88 (s, 2 H), 3.48-3.91 (m, 4H), 5.32- 5.41 (m, 1 H), 6.56 (d, J = 3.51 Hz, 1 H), 7.41 (s, 1 H), 7.46(d, J = 3.51 Hz, 1 H), 8.07 (d, J = 1.95 Hz, 1 H), 8.27 (d, J = 1.76 Hz,1 H) 2A-27 CH(CH₃)₂ H

+APCI (M + H) 407.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 1.43 (d, J = 6.64 Hz,6 H), 2.57 (s, 2 H), 2.78 (s, 2 H), 3.47 (d, J = 5.28 Hz, 1 H), 3.53 (s,2 H), 5.31- 5.40 (m, 1 H), 6.84 (d, J = 8.01 Hz, 2 H), 7.28 (d, J = 8.40Hz, 1 H), 7.36 (s, 1 H), 8.00 (s, 1 H) 2A-28 C(CH₃)₃ H

+APCI (M + H) 406.3; ¹H NMR (400 MHz, CDCl₃) δ ppm 1.54- 1.59 (m, 4 H),1.61-1.66 (m, 9 H), 2.61 (s, 2 H), 2.81 (s, 2 H), 3.46 (m, 4 H), 7.29(s, 1 H), 7.43 (dt, J = 8.60, 1.37 Hz, 1 H), 7.49 (d, J = 8.60 Hz, 1 H),7.82 (s, 1 H), 8.08-8.12 (m, 1 H) 2A-29 C(CH₃)₃ H

+ESI (M + H) 422.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 1.58 (s, 4 H), 1.63(s, 9 H), 2.60 (s, 2 H), 2.80 (s, 2 H), 3.30-3.92 (m, 4 H), 6.98 (d, J =7.82 Hz, 1 H), 7.06 (dt, J = 8.01, 1.37 Hz, 1 H), 7.08-7.12 (m, 1 H),7.29 (s, 1 H) 2A-30 C(CH₃)₃ H

+ESI MS (M + H) 421.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.96 (brs, 1 H)7.32 (s, 1 H) 7.31(d, J = 0.87 Hz, 1 H) 7.28 (dd, J = 8.00, 1.78 Hz, 1H) 6.92 (d, J = 8.00 Hz, 1 H) 3.55 (s, 2 H) 3.29 (m(5), J = 1.67 Hz, 2H) 2.98 (d., J = 0.78 Hz, 1 H) 2.88 (s, 2 H) 2.85 (d., J = 0.78 Hz, 1H), 2.64 (s, 2 H), 1.64 (s, 9 H), 1.56 (brm, 2 H) 2A-31 C(CH₃)₃ H

+ESI MS (M + H) 433.5; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.33 (s, 1 H) 7.75(s, 2 H) 7.34 (s, 1 H) 7.24 (d, J = 7.23 Hz, 1 H) 6.71 (d, J = 7.04 Hz,1 H) 3.90 (br. s., 1 H) 3.83-3.95 (m, 1 H) 3.70 (br. s., 1 H) 3.49 (br.s., 2 H) 2.91 (s, 2 H) 2.67 (br. s., 2 H) 1.66-1.72 (m, 2 H) 1.63 (s, 9H) 1.53-1.60 (m, 2 H) 2A-32 C(CH₃)₃ H

+ESI (M + H) 406.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 1.62 (br. s., 13 H),2.59 (br. s., 2 H), 2.80 (br. s., 2 H), 3.30-3.95 (m, 4 H), 7.19 (d, J =8.00 Hz, 1 H), 7.28 (s, 1 H), 7.45 (d, J = 7.61 Hz, 1 H), 7.59 (br. s.,1 H), 7.85 (s, 1 H) 2A-33 CH(CH₃)₂ H

+ESI MS (M + H) 408.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 10.77 (s, 1 H)10.70 (s, 1 H) 7.45 (s, 1 H) 6.86-7.02 (m, 3 H) 5.27 (quin, J = 6.60 Hz,1 H) 3.45 (br. s., 4 H) 3.07-3.14 (m, 1 H) 2.79 (s, 2 H) 2.60 (s, 2 H)1.48 (br. s., 4 H) 1.35 (d, J = 6.64 Hz, 6 H) 2A-34 C(CH₃)₃ H

+ESI MS (M + H) 435.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 10.15 (s, 1 H)7.39 (s, 1 H) 7.20 (d, J = 7.62 Hz, 1 H) 6.91 (dd, J = 7.62, 1.56 Hz, 1H) 6.85 (d, J = 1.56 Hz, 1 H) 3.69 (br. s., 1 H) 3.48 (br. s., 1 H)3.05-3.17 (m, 1 H) 2.89 (t, J = 7.62 Hz, 2 H) 2.82 (s, 2 H) 2.62 (s, 2H) 2.45 (dd, J = 8.70, 6.55 Hz, 2 H) 1.57 (s, 9 H) 1.46 (br. s., 4 H);ESI+ MS (M + H) 435.5. 2A-35 CH(CH₃)₂ H

+ESI MS (M + H) 421.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 10.15 (s, 1 H)7.45 (s, 1 H) 7.20 (d, J = 7.61 Hz, 1 H) 6.91 (dd, J = 7.51, 1.66 Hz, 1H) 6.85 (d, J = 1.56 Hz, 1 H) 5.27 (quin, J = 6.58 Hz, 1 H) 3.65 (br.s., 1 H) 3.53 (br. s., 1 H) 2.89 (t, J = 7.61 Hz, 2 H) 2.79 (br. s., 2H) 2.60 (s, 2 H) 2.45 (dd, J = 8.78, 6.44 Hz, 3 H) 1.48 (br. s., 4 H)1.36 (d, J = 6.63 Hz, 6 H) 2A-36 CH(CH₃)₂ H

+ESI MS (M + H) 410.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.94 (s, 1 H) 7.40(s, 1 H) 7.28-7.38 (m, 1 H) 7.00 (d, J = 10.36 Hz, 1 H) 5.38 (quin, J =6.64 Hz, 1 H) 3.26-4.02 (m, 4 H) 2.83 (s, 2 H) 2.61 (s, 2 H) 1.70 (br.s., 4 H) 1.46 (d, J = 6.45 Hz, 6 H) 2A-37 CH(CH₃)₂ H

+ESI (M = 1) 410.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 9.18 (s, 1 H) 8.56 (d,J = 6.24 Hz, 1 H) 8.15 (s, 1 H) 7.69 (d, J = 6.44 Hz, 1 H) 7.40 (s, 1 H)5.37 (dt, J = 13.27, 6.63 Hz, 1 H) 3.54-3.88 (m, 2 H) 3.17 (qd, J =7.28, 3.90 Hz, 2 H) 2.86 (s, 2 H) 2.62 (s, 2 H) 1.57-1.83 (m, 4 H) 2A-38CH(CH₃)₂ H

+ESI MS (M + H) 419.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 9.18 (s, 1 H) 8.56(d, J = 6.24 Hz, 1 H) 8.15 (s, 1 H) 7.69 (d, J = 6.44 Hz, 1 H) 7.40 (s,1 H) 5.37 (m, 6.63 Hz, 1 H) 3.54-3.88 (m, 2 H) 3.17 (qd, J = 7.28, 3.90Hz, 2 H) 2.86 (s, 2 H) 2.62 (s, 2 H) 1.57- 1.83 (m, 4 H) 2A-39 C(CH₃)₃ H

+ES1 MS (M + H) 433.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.80- 7.88 (m, 1 H)7.59-7.71 (m, 2 H) 7.40-7.49 (m, 1 H) 7.33 (s, 1 H) 5.78 (br. s., 2 H)3.87-3.98 (m, 1 H), 3.76-3.86 (m, 1 H) 3.60-3.75 (m, 2 H) 2.87 (s, 2 H),2.66 (s, 2 H) 1.68-1.79 (m, 4 H) 2A-40 CH(CH₃)₂ H

+ESI MS (M + H) 408.0; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.15 (s, 1 H) 8.09(s, 1 H) 7.42 (s, 1 H) 6.45 (s, 1 H) 5.26-5.45 (m, 1 H) 3.58-3.77 (m, 4H) 2.88 (s, 2 H) 2.65 (s, 2 H) 1.61-1.67 (m, 4 H) 1.42 (d, J = 6.64 Hz,6 H) 2A-41 C(CH₃)₃ H

ESI + MS (M + H) 407.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 13.84 (s, 1 H)8.55 (d, J = 1.95 Hz, 1 H) 8.30 (d, J = 1.37 Hz, 1 H) 8.21 (d, J = 1.17Hz, 1 H) 7.40 (s, 1 H) 3.75 (br. s., 1 H) 3.44 (br. s., 4 H) 3.05-3.17(m, 3 H) 2.84 (s, 2 H) 2.66 (s, 2 H) 1.57 (s, 9 H) 1.50 (br. s., 4 H)2A-42

H

+ESI MS (M + H) 420.2; ¹H NMR (400 MHz, CD₃OD) δ ppm 1.54 (br. s., 4 H),2.56 (s, 3 H), 2.62 (s, 2 H), 2.89 (s, 2 H), 3.38- 3.90 (m, 4 H),4.95-5.04 (m, 4 H), 6.10-6.18 (m, 1 H), 7.24 (dd, J = 8.29, 1.46 Hz, 1H), 7.52 (d, J = 8.19 Hz, 1 H), 7.54 (s, 1 H), 8.49 (s, 1 H) 2A-43C(CH₃)₃ H

ESI + MS (M + H) 422.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 8.15 (d, J =2.15 Hz, 1 H) 8.10 (s, 1 H) 7.85 (s, 2 H) 7.40 (s, 1 H) 6.46 (d, J =2.35 Hz, 1 H) 3.42- 3.65 (m, 4 H) 2.82 (s, 2 H) 2.63 (s, 2 H) 1.58 (s, 9H) 1.48-1.55 (m, 4 H) 2A-44 C(CH₃)₃ H

+ESI MS (M + H) 424.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 8.15 (d, J = 3.32Hz, 1 H) 7.62 (d, J = 0.98 Hz, 1 H) 7.32 (s, 1 H) 7.19 (dd, J = 10.75,0.98 Hz, 1 H) 3.64 (br. s., 4 H) 2.84 (s, 2 H) 2.64 (s, 2 H) 1.66 (s, 4H) 1.59 (s, 9 H) 2A-45 C(CH₃)₃ H

+ESI MS (M + H) 436.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 10.10 (s, 1 H) 7.27(s, 1 H) 7.14 (s, 1 H) 7.12 (dd, J = 7.91, 1.27 Hz, 1 H) 6.92 (d, J =8.01 Hz, 1 H) 3.64 (br. s., 4 H) 3.39 (s, 3 H) 2.78 (s, 2 H) 2.58 (s, 2H) 1.62 (s, 9 H) 1.55 (br. s., 4 H) 2A-46 CH(CH₃)₂ H

+ESI MS (M + H) 422.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 9.82 (s, 1 H) 7.36(s, 1 H) 7.15 (s, 1 H) 7.13 (dd, 1 H) 6.93 (d, J = 8.01 Hz, 1 H) 5.35(m, 1 H) 3.58 (br. s., 4 H) 3.40 (s, 3 H) 2.78 (s, 2 H) 2.57 (s, 2 H)1.58 (br. s., 4 H) 1.43 (d, J = 6.64 Hz, 6 H) 2A-47 CH(CH₃)₂ H

+ESI MS (M + H) 426.4; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.30 (s, 1 H) 7.58(br. s., 1 H) 7.38 (s, 1 H) 7.34 (s, 1H) 5.34 (m, 1 H) 3.83 (br. 5., 1H) 3.65 (br. s., 1 H) 3.46 (br. s., 2 H) 2.85 (s, 2 H) 2.61 (s, 2 H)1.43-1.73 (m, 4 H) 1.38 (d, J = 6.63 Hz, 6 H) 2A-48 C(CH₃)₃ H

+ESI MS (M + H) 440.4; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.30 (s, 1 H) 7.59(br. s., 1 H) 7.34 (s, 1 H) 7.30 (s, 1 H) 3.81 (br. s., 1 H) 3.65 (br.s., 1 H) 3.47 (br. s., 2 H) 2.86 (s, 2 H) 2.62 (s, 2 H) 1.62-1.75 (m, 2H) 1.59 (s, 9 H) 1.49-1.56 (m, 2 H) 2A-49 C(CH₃)₃ H

+ESI MS (M + H) 436.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 8.36 (s, 1 H) 7.29(s, 1 H) 7.06-7.10 (m, 2 H) 7.00 (d, 1 H) 3.57 (br. s., 4 H) 3.40 (s, 3H) 2.80 (s, 2 H) 2.60 (s, 2 H) 1.63 (s, 9 H) 1.58 (br. s., 4 H) 2A-50CH(CH₃)₂ H

+ESI MS (M + H) 422.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 9.55 (br. s., 1 H)7.36 (s, 1 H) 7.00- 7.10 (m, 3 H) 5.35 (quin, J = 6.55 Hz, 1 H) 3.58(br. s., 4 H) 3.41 (s, 3 H) 2.79 (s, 2 H) 2.58 (s, 2 H) 1.59 (br. s., 4H) 1.43 (d, J = 6.45 Hz, 6 H) 2A-51 C(CH₃)₃ H

+ESI MS(M + H) 424.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.99 (s, 1 H) 7.39(br. s., 1 H) 7.30 (s, 1 H) 7.01 (d, J = 12.12 Hz, 1 H) 3.41-3.94 (m, 4H) 2.83 (s, 1 H) 2.82 (s, 2 H) 2.61 (s, 2 H) 1.47- 1.78(m, 13 H) 2A-52CH(CH₃)₂ H

+ESI MS (M + H) 440.4; ¹H NMR (400 MHz, CD₃OD) δ ppm 7.43- 7.55 (m, 1 H)7.42 (s, 1 H) 7.30 (s, 1 H) 5.30-5.43 (m, 1 H) 3.86 (br. s., 1 H) 3.68(br. s., 1 H) 3.51 (br. s., 2 H) 2.89 (s, 2 H) 2.65 (s, 2 H) 2.61 (s, 3H) 1.52- 1.73 (m, 4 H) 1.42 (d, J = 6.64 Hz, 6 H) 2A-53 C(CH₃)₃ H

+ESI MS (M + H) 454.4; ¹H NMR (400 MHz, CD₃OD) δ ppm 7.49 (s, 1 H) 7.34(s, 1 H) 7.30 (d, J = 1.37 Hz, 1 H) 3.40-3.97 (m, 4 H) 2.90 (s, 2 H)2.66 (s, 2H) 2.61 (s, 3 H) 1.54-1.71 (m, 13 H) 2A-54 CH(CH₃)₂ H

+ESI MS (M + H) 424.4; ¹H NMR (400 MHz, CD₃OD) δ ppm 7.42 (s, 1 H) 7.36(br. s., 1 H) 7.03 (d, J = 10.36 Hz, 1 H) 5.29-5.45 (m, 1 H) 3.38-4.07(m, 4 H) 2.88 (s, 2 H) 2.64 (br. s., 2 H) 2.59 (s, 3 H) 1.48-1.82 (m, 4H) 1.42 (d, J = 6.64 Hz, 6 H) 2A-55 C(CH₃)₃ H

+ESI MS (M + H) 438.3; ¹H NMR (400 MHz, CD₃OD) δ ppm 7.35 (s, 2 H) 7.05(d, J = 10.36 Hz, 1 H) 3.38-4.10 (m, 4 H) 2.92 (s, 2 H) 2.68 (s, 2 H)2.61 (s, 3 H) 1.65 (s, 13 H) 2A-56 C(CH₃)₃ H

+ESI MS (M + H) 456.4; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 7.39 (s, 1 H),6.97 (s, 1 H) 6.84 (s, 1 H), 3.03-3.76 (m, 4 H) 2.82 (s, 2 H) 2.62 (s, 2H) 1.58 (s, 9 H) 2A-57 CH(CH₃)₂ H

+APCI MS (M + H) 442.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 7.38 (s, 1 H)7.07-7.26 (m, 1 H) 7.03 (br. s., 1 H) 5.31-5.45 (m, 1 H) 3.25-3.92 (m, 4H) 2.80 (s, 2 H), 2.60 (s, 2 H) 1.50-1.77 m, 4H) 2A-58 CH(CH₃)₂ H

ESI + MS (M + H) 422.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 8.18 (s, 1 H)7.98 (s, 1 H) 7.46 (s, 1 H) 6.98 (br. s., 2 H) 5.27 (quin, J = 6.63 Hz,1 H) 3.90 (s, 3 H) 3.48 (br. s., 4 H) 2.79 (s, 2 H) 2.60 (s, 2 H)1.46-1.56 (m, 4 H) 1.35 (d, J = 6.7 Hz, 6 H) 2A-59 C(CH₃)₃ H

ESI + MS (M + H) 436.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 8.18 (s, 1 H)7.98 (s, 1 H) 7.40 (s, 1 H) 6.98 (br. s., 2 H) 3.90 (s, 3 H) 3.38-3.62(m, 4 H) 2.81 (s, 2 H) 2.62 (s, 2 H) 1.58 (s, 9 H) 1.45-1.52 (m, 4 H)2A-60 CH(CH₃)₂ H

ESI + MS (M + H) 435.5; ¹HNMR (400 MHz, CD₃OD) δ 8.64 (d, J = 5.2 Hz,1H), 8.05 (s, 1H), 7.46 (d, J = 4.8 Hz, 1H), 7.23 (s, 1H), 5.17 (m, 1H),3.69 (m, 1H), 3.55 (m, 1H), 3.22 (m, 1H), 3.05 (m, 1H), 2.70 (s, 2H),2.45 (s, 2H), 2.29 (s, 3H), 1.51 (m, 2H), 1.39 (m, 2H), 1.20 (m, 6H)2A-61 CH(CH₃)₂ H

ESI + MS (M + Na) 426.4; ¹HNMR (400 MHz, CD₃OD) δ 9.09 (s, 1H), 8.13 (m,2H), 7.93 (m, 2H), 7.46 (s, 1H), 5.38 (m, 1H), 3.97 (m, 1H), 3.82 (m,1H), 3.68 (m, 2H), 2.95 (s, 2H), 2.70 (s, 2H), 1.70-1.80 (m, 4H), 1.43(m, 6H) 2A-62 CH(CH₃)₂ H

ESI + MS (M + H) 410.6; ¹HNMR (400 MHz, CD₃OD) δ 9.16 (d, J = 8.4 Hz,1H), 7.84 (t, 1H), 7.43 (s, 1H), 7.25 (d, J = 7.6 Hz, 1H), 5.36 (m, 1H),3.83 (m, 1H), 3.66 (m, 1H), 3.48 (m, 2H), 2.89 (s, 2H), 2.65 (s, 2H),2.16 (s, 3H), 1.71-1.62 (m, 4H), 1.42 (m, 6H) 2A-63 CH(CH₃)₂ H

ESI + MS (M + H) 404.3; ¹HNMR (400 MHz, CD₃OD) δ 9.43 (s, 1H), 8.78 (d,J = 6 Hz, 1H), 8.72 (d, J = 8.8 Hz, 1H), 8.01 (d, J = 6 Hz, 1H), 7.85(d, J = 8.4 Hz, 1H), 7.46 (s, 1H), 5.39 (m, 1H), 3.96 (m, 1H), 3.81 (m,1H), 3.49 (m, 2H), 2.94 (s, 2H), 2.69 (s, 2H), 1.78-1.65 m, 4H , 1.42 m,6H 2A-64 CH(CH₃)₂ H

ESI + MS (M + H) 392.4; ¹HNMR (400 MHz, CD₃OD) δ 8.89 (s, 1H), 8.18 (d,J = 6 Hz, 1H), 7.85 (d, J = 6 Hz, 1H), 8.01 (d, J = 6 Hz, 1H), 7.46 (s,1H), 6.99 (s, 1H), 5.39 (m, 1H), 3.80-3.89 (m, 2H), 3.22 (m, 1H), 2.94(m, 2H), 2.68 (m, 2H), 1.71-1.83 (m, 4H), 1.44 (d, J = 6.8 Hz, 6H) 2A-65CH(CH₃)₂ H

ESI + MS (M + H) 426.2; ¹HNMR (400 MHz, CD₃OD) δ 8.18 (s, 1H), 7.57 (s,1H), 7.45 (s, 1H), 7.23 (s, 1H), 5.37 (m, 1H), 3.90 (m, 1H), 3.75 (m,1H), 3.49 (m, 1H), 3.22 (m, 1H), 2.92 (s, 2H), 2.68 (s, 2H), 1.59-1.81(m, 4H), 1.30 (m, 6H) 2A-66 CH(CH₃)₂ H

ESI + MS (M + H) 439.5; ¹HNMR (400 MHz, CD₃OD) δ 8.47 (s, 2H), 7.45 (m,1H), 5.39 (m, 1H), 3.65-3.88 (m, 10 H), 3.15-3.25 (m, 2H), 2.91 (s, 2H),2.67 (s, 2H), 1.65-1.80 (m, 4H), 1.44 (d, J = 6.8 Hz, 6H 2A-67 CH(CH₃)₂H

ESI + MS (M + H) 381.7; ¹HNMR (400 MHz, CD₃OD) δ 7.38 (s, 1H), 7.34 (s,1H), 7.10 (s, 1H), 5.97 (s, 1H), 5.31 (m, 1 H), 3.91 (m, 1H), 3.82 (m,2H), 3.65 (m, 1H), 2.84 (s, 2H), 2.59 (s, 2H), 1.58 (m, 4H), 1.37 (d, J= 6.8 Hz, 6H) 2A-68 CH(CH₃)₂ H

ESI + MS (M + H) 392.5; ¹HNMR (400 MHz, CD₃OD) δ 8.16 (s, 1H), 7.92 (s,1H), 7.62 (d, J = 8.8 Hz, 1H), 7.46 (d, J = 8.8 Hz, 1H), 7.45 (s, 1H),5.39 (m, 1 H), 3.51-3.99 (m, 4H), 2.92 (s, 2H), 2.68 (s, 2H), 1.61-1.70(m, 4H), 1.43 (d, J = 6.4 Hz, 6H 2A-69 CH(CH₃)₂ H

ESI + MS (M + H) 419.5; ¹HNMR (400 MHz, CD₃OD) δ 8.99 (d, J = 4 Hz, 1H),8.86 (d, J = 7.6 Hz, 1H), 7.56-7.72 (m, 3H), 7.56 (s, 1H), 5.51 (m, 1H), 3.77-3.83 (m, 4H), 3.02 (s, 2H), 2.78 (s, 2H), 1.79 (m, 4H), 1.54(d, J = 6.8 Hz, 6H) 2A-70 CH(CH₃)₂ H

ESI + MS (M + H) 419.5; ¹HNMR (400 MHz, CD₃OD) δ 8.99 (d, J = 4 Hz, 1H),8.86 (d, J = 7.6 Hz, 1H), 7.56-7.72 (m, 3H), 7.56 (s, 1H), 5.51 (m, 1H), 3.77-3.83 (m, 4H), 3.02 (s, 2H), 2.78 (s, 2H), 1.79 (m, 4H), 1.54(d, J = 6.8 Hz, 6H) 2A-71 CH(CH₃)₂ H

ESI + MS (M + H) 392.4; ¹HNMR (400 MHz, CD₃OD) δ 8.30 (d, J = 4 Hz, 1H),8.19 (d, J = 8 Hz, 1H), 7.79 (s, 1H), 7.45 (s, 1H), 7.25 (m, 1H), 5.40(m, 1 H), 3.73-3.88 (m, 4H), 2.93 (s, 2H), 2.69 (s, 2H), 1.67 (m, 4H),1.44 (d, J = 6.4 Hz, 6H) 2A-72 CH(CH₃)₂ H

ESI + MS (M + H) 419.4; ¹HNMR (400 MHz, CD₃OD) δ 9.21 (s, 1H), 8.48 (d,J = 6.4 Hz, 1H), 8.17 (d, 1H), 7.58 (m, 1H), 7.49 (d, J = 8 Hz, 1H),7.45 (s, 1H), 5.40 (m, 1 H), 3.24-3.80 (m, 4H), 2.91 (s, 2H), 2.67 (s,2H), 1.68 (m, 4H), 1.43 (d, J = 6.4 Hz, 6H) 2A-73 CH(CH₃)₂ H

ESI + MS (M + 23) 419.4; ¹HNMR (400 MHz, CD3OD) δ 9.00 (d, J = 4.8 Hz,1H), 8.75 (d, J = 7.2 Hz, 1H), 7.857 (m, 1H), 7.58 (d, J = 9.2 Hz, 1H),7.59 (d, J = 8 Hz, 1H), 7.45 (s, 1H), 5.40 (m, 1 H), 3.58 (m, 4H), 2.92(s, 2H), 2.68 (s, 2H), 1.69 (m, 4H), 1.43 (d, J = 6.4 Hz, 6H) 2A-74CH(CH₃)₂ H

+ESI MS (M + H) 418.6; ¹HNMR (400 MHz, CD₃OD) δ 8.00 (s, 2H), 7.67 (d, J= 8 Hz, 1H), 7.61 (s, 1H), 7.41 (m, 2H), 7.22 (d, J = 7.6 Hz, 1H), 5.36(m, 1 H), 3.48-3.90 (m, 2H), 3.30 (m, 2H), 2.88 (s, 2H), 2.64 (s, 2H),1.56- 1.68 (m, 4H), 1.40 (d, J = 4.4 Hz, 6H) 2A-75 CH(CH₃)₂ CH₃

+ESI MS (M + H) 406.6; ¹H NMR (400 MHz, CDCl₃) δ ppm 10.28 (br, s., 1 H)8.12 (s, 1 H) 7.85 (s, 1 H) 7.41-7.61 (m, 2 H) 5.21-5.50 (m, 1 H)3.29-4.02 (m, 4 H) 2.69 (s, 2 H) 2.58 (s, 2 H) 2.24 (s, 3 H) 1.49-1.77(m, 4 H) 1.44 (d, J = 6.64 Hz, 6H) 2A-76 CH(CH₃)₂ CN

+ESI MS (M + H) 417.5; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.11 (s, 1 H) 7.89(d, J = 0.78 Hz, 1 H) 7.59 (dd, J = 8.60, 0.78 Hz, 1 H) 7.44 (dd, J =8.60, 0.78 Hz, 1 H) 5.42 (dt, J = 13.24, 6.57 Hz, 1 H) 3.79 (br. s., 2H) 3.54 (br. s., 2 H) 2.96 (s, 2 H) 2.71 (s, 2 H) 1.48-1.81 (m, 4 H)1.44 (d, J = 6.64 Hz, 6 H) 2A-77 C(CH₃)₃ CN

+ESI MS (M + H) 431.5; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.26 (s, 1 H) 7.68(br. s., 2 H) 7.34 (d, J = 8.60 Hz, 1 H) 3.80 (br. s., 2 H) 3.52 (br.s., 2 H) 2.98 (s, 2 H) 2.74 (s, 2 H) 1.46-1.80 (m, 13 H) 2A-78 C(CH₃)₃CN

+ESI MS (M + H) 431.5; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.11 (d, J = 0.98Hz, 1 H) 7.89 (dd, J = 1.46, 0.88 Hz, 1 H) 7.59 (dt, J = 8.58, 0.88 Hz,1 H) 7.44 (dd, J = 8.68, 1.46 Hz, 1 H) 3.78 (br. s., 2 H) 3.54 (br. s.,2 H) 2.98 (s, 2 H) 2.74 (s, 2 H) 1.52-1.82 (m, 13 H)

Example 3 Preparation of1-isopropyl-1′-(2-methyl-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(3A-1)

2-Methyl-1H-benzimidazole-5-carboxylic acid (15 g) was taken up intetrahydrofuran (500 mL), dimethylformamide (329 uL) and oxalyl chloride(22.1 mL) were added. The reaction solution was stirred at ambienttemperature for 16 hours. The solution was concentrated in vacuo and theresidue was taken up in dichloromethane and concentrated (×2) underreduced pressure. To the resulting acid chloride was addedtetrahydrofuran (500 mL), Preparation I-1A-1e (25.9 g) and triethylamine(71.2 mL). The solution was stirred at room temperature for 16 hours. Tothe reaction was added saturated, aqueous sodium bicarbonate (250 mL)and the solution was stirred for 5 min. The layers were separated andthe aqueous layer was extracted with 1:1 ethyl acetate/tetrahydrofuran.The organic layers were combined, diluted with ethyl acetate (1 L) andwashed with saturated aqueous, sodium bicarbonate (200 mL) andsaturated, aqueous sodium chloride. The organic layer was dried oversodium sulfate, filtered and concentrated in vacuo to a light yellowsolid. The solid was dissolved in hot methanol (300 mL) and then heatedto reflux. To the solution was added 350 mL ethyl acetate and 300 mL ofsolvent was removed by distillation. Additional ethyl acetate was addeddropwise until an internal temperature of 70° C. was reached. Thesolution was cooled to room temperature over 3 hours. The solids werecollected by filtration and dried in a vacuum oven (40° C.) for 16 hoursto afford the title compound as a white solid (20.5 g, 59%): +ESI MS(M+H) 406.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm 12.25-12.33 (m, 1H),7.35-7.51 (m, 3H), 7.05-7.16 (m, 1H), 5.16-5.31 (m, 1H), 3.32-3.58 (m,4H), 2.77 (s, 2H), 2.57 (s, 2H), 1.40-1.52 (m, 4H), 1.32 (d, J=6.45 Hz,6H).

In the present example it is to be understood that the starting material2-Methyl-1H-benzimidazole-5-carboxylic acid employed in this examplealso exists as its tautomeric form2-Methyl-1H-benzimidazole-6-carboxylic acid (also known as2-Methyl-3H-benzimidazole-5-carboxylic acid) and each is designated bythe same CAS No. 709-19-3. It is to be further understood that theinstant example has been depicted above as one of two tautomeric formsof the compound with respect to the 2-methyl benzimidazolyl group andthat the title compound is synonomous with the tautomeric form1-isopropyl-1′-(2-methyl-1H-benzo[d]imidazole-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-onewhich is depicted as:

Example 4 Preparation of1′-(1H-indazole-5-carbonyl)-1-isopropyl-6-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(4A-1)

Preparation I-4A-1e (177 mg, 0.677 mmol), 1H-indazole-5-carboxylic acid(110 mg, 0.677 mmol),O-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (257 mg, 0.677 mmol) and triethylamine (138 mg, 1.35mmol) were combined in 3 mL of dimethylformamide and stirred at roomtemperature for 18 hours. To the reaction was added saturated, aqueoussodium bicarbonate (2 mL). The reaction was partitioned between ethylacetate (80 mL) and water (20 mL). The phases were separated and theaqueous layer was extracted with ethyl acetate (50 mL). The organicphases were combined, dried over magnesium sulfate and concentrated togive an oil. The oil was purified by flash chromatography using 0-5%methanol in dichloromethane as eluent to afford the title compound as asolid (196 mg, 72%): +APCI MS (M+H) 406.2; ¹H NMR (400 MHz, DMSO-d₆) δppm 13.21 (br. s., 1H), 8.13 (s, 1H), 7.82 (s, 1H), 7.56 (d, J=8.60 Hz,1H), 7.46 (s, 1H), 7.37 (dd, J=8.60, 1.37 Hz, 1H), 5.25 (m, 1H),3.21-3.39 (m, 4H), 3.01-3.19 (m, 1H), 2.74-2.94 (m, 2H), 1.41-1.62 (m,4H), 1.34 (d, J=6.64 Hz, 6H), 1.07 (d, J=7.23 Hz, 3H).

Example 5 Preparation of(+)-1′-(1H-indazole-5-carbonyl)-1-isopropyl-6-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(5A-1) and(−)-1′-(1H-indazole-5-carbonyl)-1-isopropyl-6-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(5A-2)

Racemic compound Example 4 (Compound 4A-1) was separated to give thecorresponding two enantiomers using chiral HPLC: [Chiralpakl AD-H(10×250); mobile phase: 70:30 (CO₂/Ethanol); flow rate=10 mL/min].Compound 5A-1: retention time=4.17 min; optical rotation results:c=0.0053 g/mL in ethanol; path length=1 dcm; observed rotation=+0.202 (Dline of a sodium lamp (589 nm) at 20° C.); specific rotation=+38.1.Compound 5A-2: retention time=5.47 min; optical rotation results:c=0.0053 g/mL in ethanol; path length=1 dcm; observed rotation=−0.184 (Dline of a sodium lamp (589 nm) at 20° C.); specific rotation=−34.1.

Example 6 Preparation of1′-(1H-indazole-5-carbonyl)-1-isopropyl-6,6-dimethyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(6A-1)

A solution of 1H-indazole-5-carboxylic acid (27 mg, 0.167 mmol) indimethylformamide (2 mL) was treated with N-methyl morpholine (51 mg,0.167 mmol) followed by 2-chloro-4,6-dimethoxy-1,3,5-triazine (29 mg,167 mmol). The solution was stirred at ambient temperature for twohours. To the reaction was added Preparation I-6A-1b (46 mg, 0.17 mmol)and N-methyl morpholine (34 mg, 0.334 mmol) as a solution indimethylformamide (2 mL). The reaction mixture was stirred at roomtemperature for 18 hours. To the reaction was added aqueous, saturatedammonium chloride (1 mL). The reaction was partitioned between ethylacetate (30 mL) and water (5 mL). The phases were separated and theaqueous layer was extracted with ethyl acetate (30 mL). The organicphases were combined, dried over magnesium sulfate and concentrated togive an oil. The oil was purified by flash chromatography using 0-5%methanol in dichloromethane as eluent to afford the title compound as asolid (39 mg, 56%): +APCI MS (M+H) 420.3; ¹H NMR (400 MHz, DMSO-d₆) δppm 13.18 (s, 1H), 8.10 (s, 1H), 7.80 (s, 1H), 7.53 (d, J=8.78 Hz, 1H),7.42 (s, 1H), 7.34 (dd, J=8.58, 1.37 Hz, 1H), 5.24 (m, 1H), 3.84 (br.s., 6H), 1.38-1.72 (m, 4H), 1.32 (d, J=6.63 Hz, 6H), 1.06 (s, 6H).

Example 7 Preparation of1′-(1H-benzo[d]imidazole-5-carbonyl)-1-isopropyl-3-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(7A-1)

Preparation I-2A-75f (150 mg, 0.50 mmol),1H-benzo[d]imidazole-5-carboxylic acid (82 mg, 0.50 mmol),O-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (198 mg, 0.50 mmol) and triethylamine (103 mg, 1.01mmol) were combined in 3 mL of dimethylformamide and stirred at roomtemperature for 16 hours. The reaction was partitioned between 10 mLethyl acetate and 10 mL saturated aqueous sodium bicarbonate. Theorganic phase was separated and then concentrated to give an oil. Theoil was purified by flash chromatography using 5-10% ethanol indichloromethane as eluent to afford the title compound as a solid (82mg, 40%): +APCI MS (M+H) 406.2; ¹H NMR (400 MHz, CDCl₃) δ ppm 8.05 (s,1H), 7.71 (s, 1H), 7.43 (m, 2H), 5.35 (m, 1H), 3.62 (m, 4H), 2.82 (s,2H), 2.57 (br. s., 2H), 2.24 (s, 3H), 1.49-1.89 (m, 4H), 1.44 (d, J=6.64Hz, 6H).

Example 8 Preparation of1′-(1H-benzo[d]imidazole-5-carbonyl)-1-isopropyl-7-oxo-1,4,6,7-tetrahydrospiro[indazole-5,4′-piperidine]-3-carbonitrile(8A-1)

Preparation I-2A-76b (50 mg, 0.18 mmol),1H-benzo[d]imidazole-5-carboxylic acid (30 mg, 0.18 mmol),O-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (77 mg, 0.20 mmol) and triethylamine (56 mg, 0.55mmol) were combined in 2.3 mL of dichloromethane and stirred at roomtemperature for 16 hours. To the reaction was added saturated aqueoussodium bicarbonate (3 mL). The phases were separated and the aqueousphase was extracted with an additional portion of dichloromethane. Theorganic phases were combined and concentrated to give a residue. Theresidue was dissolved in methanol and potassium carbonate (49 mg, 0.36mmol) was added. The mixture was stirred at ambient temperature for 5minutes. The reaction was quenched with the addition of saturatedaqueous ammonium chloride (2 mL) and the methanol was concentrated invacuo to give a residue. The residue was partitioned between water anddichloromethane and then the phases were separated. The organic layerwas washed with saturated aqueous sodium chloride, dried over magnesiumsulfate and concentrated in vacuo to give a residue which was purifiedby flash chromatography using 0-10% methanol in ethyl acetate as eluentto afford the title compound as a solid (17 mg, 22%): +ESI MS(M+H)=417.5; ¹H NMR (400 MHz, CD₃OD) δ ppm 8.26 (s, 1H), 7.51-7.88 (m,2H), 7.33 (d, J=9.77 Hz, 1H), 5.35-5.47 (m, 1H), 3.33-3.91 (m, 4H), 2.96(s, 2H), 2.71 (s, 2H), 1.48-1.83 (m, 4H), 1.44 (d, J=6.64 Hz, 6H).

Example 9 Preparation of1′-(1H-indazole-5-carbonyl)-1-(oxetan-3-yl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(9A-1)

Preparation I-2A-42 g (12.4 mg, 0.042 mmol), 1H-indazole-5-carboxylicacid (7 mg, 0.043 mmol),O-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (16 mg, 0.042 mmol) and triethylamine (14 mg, 0.14mmol) were combined in 3 mL of dimethylformamide and stirred at roomtemperature for 16 hours. The reaction was diluted with ethyl acetateand washed with citric acid (0.5 M in water), saturated aqueous sodiumchloride and saturated aqueous sodium bicarbonate one time each. Theorganic phase was separated and then dried over magnesium sulfate,filtered and concentrated to give a residue. The residue was purified byflash chromatography using 0-20% methanol in ethyl acetate as eluent toafford the title compound as an oil (7 mg, 40%): +APCI MS (M+H) 406.2;¹H NMR (400 MHz, CD₃OD) δ ppm 8.15 (s, 1H), 7.91 (s, 1H), 7.63 (d,J=8.60 Hz, 1H), 7.58 (s, 1H), 7.46 (dd, J=8.70, 1.47 Hz, 1H), 6.10-6.25(m, 1H), 4.95-5.13 (m, 4H), 3.41-4.06 (m, 4H), 2.93 (s, 2H), 2.66 (s,2H), 1.44-1.85 (m, 4H).

Example 10 Preparation of1′-(1H-indazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(10A-1)

Preparation I-1A-1e (30.3 g, 94.6 mmol) and 1H-indazole-5-carboxylicacid (16.96 g, 104.6 mmol) were suspended in dimethyl acetamide (430 mL)and 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride (22.3g, 115 mmol) was added, followed by the dropwise addition oftriethylamine (65 mL, 475 mmol). 1-Hydroxybenzotriazole hydrate (16.2 g,106 mmol) was then added and the reaction mixture was stirred at 60° C.for 2 hours. The reaction was poured into half saturated, aqueousammonium chloride (500 mL) and extracted with ethyl acetate (1×1 L,2×500 mL). The combined organic layers were washed with aqueous sodiumbicarbonate (2×500 mL), water (3×500 mL) and aqueous saturated sodiumchloride (1×500 mL). The organic layer was dried over sodium sulfate,filtered and concentrated under reduced pressure to an oil. The oil waspurified via flash column chromatography (1-6% methanol indichloromethane) to afford the desired product (27.1 g). A small amountwas crystallized using ethyl acetate/heptane. This was used to seed thefollowing crystallization. The product was dissolved in ethyl acetate(100 mL) and heated to reflux until the solution turned hazy. A smallamount of seed crystal was added. The mixture was cooled to roomtemperature and a precipitate formed and was stirred 80 hours. Theprecipitate was collected by filtration and washed with cold ethylacetate (2×30 mL). The material was air dried and then further driedunder high vacuum to afford the desired title product as an off-whitesolid (23 g, 62%). +ESI MS (M+H) 392.5; ¹H NMR (400 MHz, DMSO-d₆) δ ppm13.19 (s, 1H), 8.08-8.12 (m, 1H), 7.78-7.80 (m, 1H), 7.49-7.57 (m, 1H),7.43 (s, 1H), 7.29-7.38 (m, 1H), 5.17-5.31 (m, 1H), 3.45 (br. s., 4H),2.78 (s, 2H), 2.59 (s, 2H), 1.48 (br. s., 4H), 1.32 (d, J=6.63 Hz, 6H).

Example 11 Preparation of1′-(4-(1H-imidazol-2-yl)benzoyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one(11A-1)

4-(1H-imidazol-2-yl)benzoic acid (1.4 mg, 75 umol), anhydrousacetonitrile (400 ul), Preparation I-1A-1f (1.9 mg, 75 ul),triethylamine (21 ul, 150 umol) andO-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (2.9 mg, 75 umol) were combined and heated to 30° C.for 14 hours. The reaction solution was concentrated in vacuo andpurified by preparative HPLC to give the title compound. PreparativeHPLC method: column=Luna 5u 100×21.2 mm, solvent phase A=0.1%trifluoroacetic acid in water, phase B=acetonitrile, flow rate=23 mL/minand detector=UV.

Gradient:

Time (min) Phase B (%) 0 20 1.5 20 8 50 linear 8.2 100 9.4 100 9.6 20 1020Analytical HPLC method: column=welch materials XB-C18 2.1*50 mm, solventphase A=H₂O (1 L H₂O with 0.5 mL NH₃.H₂O), solvent phase B=acetonitrile,flow rate: 0.8 mL/min. Retention time=2.419 minutes.Gradient:

Time (min) Phase A (%) Phase B (%) 0 95 5 0.5 95 5 3.4 0 100 4.2 0 1004.21 95 5 4.70 95 5Mass parameter: Mass range=170-1000 Fragmentor=50 Gas flow=10 L/min Drygas temperature=350° C. Capillary voltage (v)=2500 M+H=418

The compounds listed in Table 3 below were prepared using proceduresanalogous to those described above for the synthesis of Example 11,Compound 11A-1 using the appropriate starting materials which areavailable commercially, prepared using preparations well-known to thoseskilled in the art, or prepared in a manner analogous to routesdescribed above for other intermediates. The compounds listed below wereisolated initially as the free base and may be converted to theircorresponding hydrochloride salt for testing. The compounds werecharacterized by the same analytical HPLC method referenced for Example11, Compound 11A-1 (method A) or by a method in which 0.05%trifluoroacetic acid in water was substituted for solvent phase A(method B).

TABLE 3

Example R¹ C(O)R⁴ Analytical Data 11A-2 CH(CH₃)₂

MS (M + H) 396; HPLC method A = 2.504 min 11A-3 CH(CH₃)₂

MS (M + H) 418; HPLC method B = 2.490 min 11A-4 CH(CH₃)₂

MS (M + H) 433; HPLC method A = 2.649 min 11A-5 C(CH₃)₃

MS (M + H) 447; HPLC method A = 2.817 min 11A-6 CH(CH₃)₂

MS (M + H 434; HPLC method B = 3.356 min 11A-7 C(CH₃)₃

MS (M + H) 432; HPLC method B = 2.637 min 11A-8 C(CH₃)₃

MS (M + H) 447; HPLC method A = 2.560 min 11A-9 CH(CH₃)₂

MS (M + H) 417; HPLC method B = 2.983 min 11A-10 CH(CH₃)₂

MS (M + H) 396; HPLC method B = 2.584 min 11A-11 C(CH₃)₃

MS (M + H) 410; HPLC method B = 2.814 min 11A-12 C(CH₃)₃

MS (M + H) 448; HPLC method B = 2.291 min 11A-13 CH(CH₃)₂

MS (M + H) 432; HPLC method A = 2.369 min 11A-14 CH(CH₃)₂

MS (M + H) 406; HPLC method B = 2.489 min 11A-15 CH(CH₃)₂

MS (M + H) 418; HPLC method B = 2.944 min 11A-16 C(CH₃)₃

MS (M + H) 420; HPLC method A = 2.945 min 11A-17 C(CH₃)₃

MS (M + H) 432; HPLC method B = 2.781 min 11A-18 CH(CH₃)₂

MS (M + H) 419; HPLC method B = 2.609 min 11A-19 C(CH₃)₃

MS (M + H) 406; HPLC method B = 2.845 min 11A-20 C(CH₃)₃

MS (M + H) 431; HPLC method B = 3.164 min 11A-21 CH(CH₃)₂

MS (M + H) 418; HPLC method B = 3.098 min 11A-22 C(CH₃)₃

MS (M + H) 452; HPLC method A = 2.869 min 11A-23 C(CH₃)₃

MS (M + H) 432; HPLC method A = 2.949 min 11A-24 CH(CH₃)₂

MS (M + H) 438; HPLC method B = 2.578 min 11A-25 C(CH3)3

MS (M + H) 400; HPLC method B = 2.883 min 11A-26 C(CH₃)₃

MS (M + H) 433; HPLC method A = 2.696 min 11A-27 C(CH₃)₃

MS (M + H) 420; HPLC method B = 2.622 min 11A-28 CH(CH₃)₂

MS (M + H) 392; HPLC method B = 2.406 min 11A-29 C(CH₃)₃

MS (M + H) 433; HPLC method B = 2.614 min 11A-30 C(CH₃)₃

MS (M + H) 410; HPLC method B = 2.591 min 11A-31 CH(CH₃)₂

MS (M + H) 393; HPLC method A = 2.377 min 11A-32 C(CH₃)₃

MS (M + H) 406; HPLC method A = 2.679 min 11A-33 CH(CH₃)₂

MS (M + H) 393; HPLC method A = 2.245 min 11A-34 CH(CH₃)₂

MS (M + H) 392; HPLC method A = 2.694 min 11A-35 CH(CH₃)₂

MS (M + H) 409; HPLC method A = 1.947 min 11A-36 C(CH₃)₃

MS (M + H) 405; HPLC method B = 2.721 min 11A-37 CH(CH₃)₂

MS (M + H) 419; HPLC method B = 2.669 min 11A-38 C(CH₃)₃

MS (M + H) 410; HPLC method B = 2.542 min 11A-39 C(CH₃)₃

MS (M + H) 407; HPLC method A = 2.436 min 11A-40 CH(CH₃)₂

MS (M + H) 406; HPLC method B = 2.431 min 11A-41 CH(CH₃)₂

MS (M + H) 418; HPLC method B = 2.424 min 11A-42 CH(CH₃)₂

MS (M + H) 438; HPLC method A = 2.595 min 11A-43 CH(CH₃)₂

MS (M + H) 391; HPLC method A = 2.909 min 11A-44 C(CH₃)₃

MS (M + H) 397; HPLC method B = 2.787 min 11A-45 C(CH₃)₃

MS (M + H) 406; HPLC method B = 2.858 min 11A-46 CH(CH₃)₂

MS (M + H) 396; HPLC method B = 2.391 min 11A-47 CH(CH₃)₂

MS (M + H) 392; HPLC method A = 2.704 min 11A-48 CH(CH₃)₂

MS (M + H) 392; HPLC method A = 2.522 min 11A-49 CH(CH₃)₂

MS (M + H) 383; HPLC method B = 2.546 min 11A-50 C(CH₃)₃

MS (M + H) 433; HPLC method A = 2.823 min 11A-51 C(CH₃)₃

MS (M + H) 406; HPLC method B = 2.554 min 11A-52 CH(CH₃)₂

MS (M + H) 392; HPLC method B = 2.309 min 11A-53 C(CH₃)₃

MS (M + H) 432; HPLC method B = 2.603 min 11A-54 CH(CH₃)₂

MS (M + H) 419; HPLC method A = 2.577 min 11A-55 CH(CH₃)₂

MS (M + H) 406; HPLC method A = 2.509 min 11A-56 CH(CH₃)₂

MS (M + H) 392; HPLC method B = 2.353 min 11A-57 C(CH₃)₃

MS (M + H) 452; HPLC method A = 2.798 min 11A-58 C(CH₃)₃

MS (M + H) 407; HPLC method A = 2.572 min 11A-59 C(CH₃)₃

MS (M + H) 433; HPLC method A = 3.026 min 11A-60 CH(CH₃)₂

MS (M + H) 392; HPLC method A = 2.480 min 11A-61 C(CH₃)₃

MS (M + H) 418; HPLC method A = 2.732 min 11A-62 CH(CH₃)₂

MS (M + H) 419; HPLC method A = 2.514 min 11A-63 CH(CH₃)₂

MS (M + H) 419; HPLC method A = 2.445 min 11A-64 C(CH₃)₃

MS (M + H) 432; HPLC method B = 2.613 min 11A-65 CH(CH₃)₂

MS (M + H) 393; HPLC method A = 2.280 min 11A-66 CH(CH₃)₂

MS (M + H) 423; HPLC method B = 2.426 min 11A-67 C(CH₃)₃

MS (M + H) 406; HPLC method A = 2.648 min 11A-68 C(CH₃)₃

MS (M + H) 407; HPLC method A = 2.471 min 11A-69 C(CH₃)₃

MS (M + H) 406; HPLC method B = 3.221 min 11A-70 C(CH₃)₃

MS (M + H) 437; HPLC method A = 2.036 min 11A-71 C(CH₃)₃

MS (M + H) 392; HPLC method A = 2.457 min 11A-72 C(CH₃)₃

MS (M + H) 433; HPLC method A = 2.776 min 11A-73 C(CH₃)₃

MS (M + H) 406; HPLC method B = 2.804 min 11A-74 C(CH₃)₃

MS (M + H) 406; HPLC method B = 2.800 min

Example 12 Preparation of1-tert-butyl-1′-(1H-pyrrolo[2,3-b]pyridine-2-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

Preparation I-1a-2f (88 mg, 0.3 mmol),1H-Pyrrolo[2,3-b]pyridine-2-carboxylic acid (48 mg, 0.3 mmol),O-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (116 mg, 0.3 mmol) and triethylamine (0.83 mL, 0.59mmol) were combined in 3 mL of dimethylformamide and stirred at roomtemperature for 30 hours. The reaction was partitioned between 10 mLethyl acetate and 10 mL saturated aqueous ammonium chloride. The organicphase was separated and then washed with saturated, aqueous sodiumbicarbonate and saturated, aqueous sodium chloride, dried over magnesiumsulfate and concentrated to give a cream-colored solid. The solid waspurified by flash chromatography using 0-20% methanol in ethyl acetateas eluent to afford the title compound as a white solid (91 mg, 76%):+ESI MS (M+H) 406.5; ¹H NMR (400 MHz, CDCl₃) δ ppm 11.05 (br. s., 1H),8.50 (dd, J=4.69, 1.56 Hz, 1H), 7.94 (dd, J=8.01, 1.56 Hz, 1H), 7.30 (s,1H), 7.11 (dd, J=8.01, 4.69 Hz, 1H), 6.67 (s, 1H), 3.91 (br. s., 2H),3.86 (br. s., 2H), 2.83 (s, 2H), 2.63 (s, 2H), 1.65 (br.s, 13H).

Example 13 Preparation of3-bromo-1′-(7-bromo-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carbonyl)-1-tert-butyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one

Preparation I-13A-1c (190 mg, 0.504 mmol),7-bromo-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carboxylic acid (130mg, 0.506 mmol), O-(azabenzotriazole-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate (205 mg, 0.52 mmol) and triethylamine (100 mg, 1.0mmol) was combined in dichloromethane (10 mL) and stirred at ambienttemperature for 16 hours. To the reaction solution was added ethylacetate (30 mL). The reaction solution was washed with 10% wt/wt aqueouscitric acid (5 mL), saturated aqueous sodium bicarbonate (5 mL) andsaturated, aqueous sodium chloride (5 mL). The organic layer was driedover magnesium sulfate and concentrated to give a residue which waspurified by silica gel chromatography with a 2-8% methanol indichloromethane gradient to give 13A-1 (52 mg, 18%): +ESI MS (M+H)580.4; ¹H NMR (400 MHz, CDCl₃) δ ppm 10.05 (br. s., 1H), 9.74 (br. s.,1H), 6.95-7.37 (m, 2H), 3.17-4.03 (m, 4H), 2.70 (br. s., 2H), 2.60 (br.s., 2H), 1.62 (br.s, 13H).

Pharmacological Data

Biological Protocols

The utility of the compounds of present invention, in the treatment ofdiseases (such as are detailed herein) in animals, particularly mammals(e.g., humans) may be demonstrated by the activity thereof inconventional assays known to one of ordinary skill in the art, includingthe in vitro and in vivo assays described below. Such assays alsoprovide a means whereby the activities of the compound of the presentinvention can be compared with the activities of other known compounds.

Direct Inhibition of the Activities of ACC1 and ACC2

The ACC inhibitory activity of the compound of the present invention wasdemonstrated by methods based on standard procedures. For example directinhibition of ACC activity, for the compound of Formula (1) wasdetermined using preparations of recombinant human ACC1 (rhACC1) andrecombinant human ACC2 (rhACC2). Representative sequences of therecombinant human ACC1 and ACC2 that can be used in the assay areprovided in FIG. 1 (SEQ ID NO. 1) and FIG. 2 (SEQ. ID NO. 2),respectively.

[1] Preparation of rhACC1. Two liters of SF9 cells, infected withrecombinant baculovirus containing full length human ACC1 cDNA, weresuspended in ice-cold lysis buffer (25 mM Tris, pH 7.5; 150 mM NaCl; 10%glycerol; 5 mM imidazole (EMD Bioscience; Gibbstown, N.J.); 2 mM TCEP(BioVectra; Charlottetown, Canada); Benzonase nuclease (10000 U/100 gcell paste; Novagen; Madison, Wis.); EDTA-free protease inhibitorcocktail (1 tab/50 mL; Roche Diagnostics; Mannheim, Germany). Cells werelysed by 3 cycles of freeze-thaw and centrifuged at 40,000×g for 40minutes (4° C.). Supernatant was directly loaded onto a HisTrap FF crudecolumn (GE Healthcare; Piscataway, N.J.) and eluted with an imidazolegradient up to 0.5 M over 20 column volumes (CV). ACC1-containingfractions were pooled and diluted 1:5 with 25 mM Tris, pH 7.5, 2 mMTCEP, 10% glycerol and direct loaded onto a CaptoQ (GE Healthcare)column and eluted with an NaCl gradient up to 1 M over 20 CV's.Phosphate groups were removed from purified ACC1 by incubation withlambda phosphatase (100 U/10 μM target protein; New England Biolabs;Beverly, Mass.) for 14 hours at 4° C.; okadaic acid was added (1 μMfinal concentration; Roche Diagnostics) to inhibit the phosphatase.Purified ACC1 was exchanged into 25 mM Tris, pH 7.5, 2 mM TCEP, 10%glycerol, 0.5 M NaCl by 6 hour dialysis at 4° C. Aliquots were preparedand frozen at −80° C.

[2] Measurement of rhACC1 inhibition. hACC1 was assayed in a Costar#3676 (Costar, Cambridge, Mass.) 384-well plate using the TranscreenerADP detection FP assay kit (Bellbrook Labs, Madison, Wis.) using themanufacturer's recommended conditions for a 50 μM ATP reaction. Thefinal conditions for the assay were 50 mM HEPES, pH 7.2, 10 mM MgCl₂,7.5 mM tripotassium citrate, 2 mM DTT, 0.1 mg/mL BSA, 30 μM acetyl-CoA,50 μM ATP, and 10 mM KHCO₃. Typically, a 10 μl reaction was run for 120min at 25° C., and 10 μl of Transcreener stop and detect buffer wasadded and the combination incubated at room temp for an additional 1hour. The data was acquired on a Envision Fluorescence reader(Perkinelmer) using a 620 excitation Cy5 FP general dual mirror, 620excitation Cy5 FP filter, 688 emission (S) and a 688 (P) emissionfilter.

[3] Preparation of rhACC2. Human ACC2 inhibition was measured usingpurified recombinant human ACC2 (hrACC2). Briefly, a full length Cytomaxclone of ACC2 was purchased from Cambridge Bioscience Limited and wassequenced and subcloned into PCDNA5 FRT TO-TOPO (Invitrogen, Carlsbad,Calif.). The ACC2 was expressed in CHO cells by tetracycline inductionand harvested in 5 liters of DMEM/F12 with glutamine, biotin, hygromycinand blasticidin with 1 μg/mL tetracycline (Invitrogen, Carlsbad,Calif.). The conditioned medium containing ACC2 was then applied to aSoftlink Soft Release Avidin column (Promega, Madison, Wis.) and elutedwith 5 mM biotin. 4 mgs of ACC2 were eluted at a concentration of 0.05mg/mL (determined by A280) with an estimated purity of 95% (determinedby A280). The purified ACC2 was dialyzed in 50 mM Tris, 200 mM NaCl, 4mM DTT, 2 mM EDTA, and 5% glycerol. The pooled protein was frozen andstored at −80° C., with no loss of activity upon thawing. Formeasurement of ACC2 activity and assessment of ACC2 inhibition, testcompounds were dissolved in DMSO and added to the rhACC2 enzyme as a 5×stock with a final DMSO concentration of 1%.

[4] Measurement of human ACC2 inhibition. hACC2 was assayed in a Costar#3676 (Costar, Cambridge, Mass.) 384-well plate using the TranscreenerADP detection FP assay kit (Bellbrook Labs, Madison, Wis.) using themanufacturer's recommended conditions for a 50 uM ATP reaction. Thefinal conditions for the assay were 50 mM HEPES, pH 7.2, 5 mM MgCl₂, 5mM tripotassium citrate, 2 mM DTT, 0.1 mg/mL BSA, 30 μM acetyl-CoA, 50μM ATP, and 8 mM KHCO₃. Typically, a 10 μl reaction was run for 50 minat 25° C., and 10 μl of Transcreener stop and detect buffer was addedand the combination incubated at room temp for an additional 1 hour. Thedata was acquired on an Envision Fluorescence reader (Perkinelmer) usinga 620 excitation Cy5 FP general dual mirror, 620 excitation Cy5 FPfilter, 688 emission (S) and a 688 (P) emission filter.

The results using the recombinant hACC1 and recombinant hACC2Transcreener assays described above are summarized in the table belowfor the Compounds of Formula (I) exemplified in the Examples above.

hACC1 hACC2 Example (nM) n (nM) n  1A-1 267 3 68 3  1A-2 444 3 62.3 3 1A-3 646 3 155 6  1A-4 2980 3 719 3  1A-5 800 3 127 3  2A-1 282 3 69.74  2A-2 8300 3 3570 3  2A-3 2720 2 680 3  2A-4 8730 3 2530 3  2A-5 12203 481 3  2A-6 640 3 88.8 4  2A-7 4340 3 965 4  2A-8 16000 2 4890 3  2A-94260 1 1850 3  2A-10 188 3 29.2 3  2A-11 261 3 33.5 3  2A-12 94 3 32.2 9 2A-13 95.3 3 28 5  2A-14 172 3 27 7  2A-15 2030 3 775 3  2A-16 46.9 313.6 7  2A-17 795 3 287 3  2A-18 1010 2 458 3  2A-19 57.4 3 22.1 9 2A-20 121 4 93 5  2A-21 93.3 2 32.7 4  2A-22 1890 1 334 3  2A-23 32.3 323.3 9  2A-24 30.7 3 15.8 5  2A-25 69.5 3 22.7 6  2A-26 43.8 3 42.4 4 2A-27 308 1 91.8 4  2A-28 47.8 3 17 5  2A-29 24.1 3 9.81 10  2A-30 1702 51 3  2A-31 43.5 3 28.8 4  2A-32 37 3 28.6 6  2A-33 53.7 3 14.5 10 2A-34 72.4 2 32.1 3  2A-35 97.3 2 34.1 4  2A-36 123 2 29.6 4  2A-3787.4 2 34.5 3  2A-38 297 2 99.7 3  2A-39 527 2 122 6  2A-40 262 2 116 3 2A-41 176 2 90 3  2A-42 1300 3 766 3  2A-43 304 2 149 3  2A-44 106 230.2 3  2A-45 73.7 2 32.9 3  2A-46 167 3 29.5 6  2A-47 48.2 3 15.9 5 2A-48 51.1 3 17.2 8  2A-49 92.1 2 71.6 3  2A-50 269 5 64.1 7  2A-5172.7 2 29.9 3  2A-52 54.2 3 19.3 7  2A-53 39.6 3 17.2 8  2A-54 214 251.5 3  2A-55 266 2 105 3  2A-56 31.2 3 15.1 4  2A-57 66.5 4 20.8 5 2A-58 280 2 74.1 3  2A-59 115 3 69.2 4  2A-60 8240 1 3990 1  2A-61 23201 2190 1  2A-62 4210 1 1280 1  2A-63 9330 1 3200 1  2A-64 229 1 253 1 2A-65 74.9 1 51.5 1  2A-66 30000 1 14000 1  2A-67 799 4 653 3  2A-6868.4 4 51.2 3  2A-69 122 4 42.9 3  2A-70 9790 4 3760 3  2A-71 1060 4 3873  2A-72 599 4 290 3  2A-73 516 4 139 3  2A-74 128 4 47.3 3  2A-75 113 337.7 6  2A-76 6620 1 1500 3  2A-77 6160 1 1300 3  2A-78 3840 1 1270 3 3A-1 96.1 8 48.6 17  4A-1 399 2 125 3  5A-1 4480 1 866 3  5A-2 175 180.7 4  6A-1 1530 1 373 3  7A-1 63.2 1 39.8 4  8A-1 7620 1 1660 3  9A-1704 3 264 3 10A-1 43.8 3 22.3 5 11A-1 147 4 31.8 4 11A-2 1480 2 658 411A-3 74.7 3 25.4 4 11A-4 19600 1 5800 3 11A-5 16400 1 13200 3 11A-6 9891 459 3 11A-7 36.8 3 10.4 3 11A-8 40.9 1 26.8 3 11A-9 34.4 3 19.7 511A-10 6670 1 789 3 11A-11 10600 1 1450 3 11A-12 152 1 850 3 11A-13 37.61 35.7 3 11A-14 418 1 253 3 11A-15 112 4 43.8 5 11A-16 621 1 190 311A-17 129 2 48.4 3 11A-18 235 2 56.3 4 11A-19 242 2 71.6 3 11A-20 25.53 8.46 3 11A-21 112 1 73.4 3 11A-22 1260 1 539 3 11A-23 87.1 3 20.4 511A-24 6060 1 836 3 11A-25 331 1 745 3 11A-26 130 2 41.5 4 11A-27 584 1309 3 11A-28 917 1 391 3 11A-29 151 1 64.5 5 11A-30 224 2 106 4 11A-31117 1 83.4 3 11A-32 155 1 96.4 3 11A-33 272 2 93.1 4 11A-34 2140 1 638 311A-35 3340 1 2130 3 11A-36 194 2 75.6 4 11A-37 1030 1 463 3 11A-38 21702 1030 4 11A-39 69 2 81.3 4 11A-40 1140 2 588 4 11A-41 122 4 30.1 511A-42 11700 1 2850 3 11A-43 143 1 45.6 3 11A-44 4170 1 4600 3 11A-451500 1 745 3 11A-46 391 2 97.1 4 11A-47 850 1 373 3 11A-48 78.9 2 41.2 411A-49 13500 1 4910 3 11A-50 581 1 371 3 11A-51 110 1 108 3 11A-52 550 1176 3 11A-53 68.3 3 22.8 5 11A-54 1500 1 295 3 11A-55 213 2 60.4 411A-56 1010 1 359 3 11A-57 9230 1 2330 3 11A-58 131 1 73.6 3 11A-59 86.81 80.1 3 11A-60 404 1 171 3 11A-61 2570 1 1350 3 11A-62 338 2 112 411A-63 8720 1 2550 3 11A-64 105 3 24.4 4 11A-65 1080 1 388 3 11A-66 40801 1390 3 11A-67 203 1 139 3 11A-68 825 2 285 4 11A-69 580 2 288 4 11A-701860 1 1040 3 11A-71 1560 1 285 3 11A-72 867 1 153 3 11A-73 37.4 1 37.43 11A-74 23.7 3 16.8 4 12A-1 39.5 3 67.3 4 13A-1 264 3 316 3

Acute In Vivo Assessment of ACC Inhibition in Experimental Animals

The ACC inhibitory activity of the compounds of the present inventioncan be confirmed in vivo by evaluation of their ability to reducemalonyl-CoA levels in liver and muscle tissue from treated animals.

Measurement of malonyl-CoA production inhibition in experimentalanimals. In this method, male Sprague-Dawley Rats, maintained onstandard chow and water ad libitum (225-275 g), were randomized prior tothe study. Animals were either fed, or fasted for 18 hours prior to thebeginning of the experiment. Two hours into the light cycle the animalswere orally dosed with a volume of 5 mL/kg, (0.5% methyl cellulose;vehicle) or with the appropriate compound (prepared in vehicle). Fedvehicle controls were included to determine baseline tissue malonyl-CoAlevels while fasted animals were included to determine the effectfasting had on malonyl-CoA levels. One hour after compoundadministration the animals were asphyxiated with CO₂ and the tissueswere removed. Specifically, blood was collected by cardiac puncture andplaced into BD Microtainer tubes containing EDTA (BD Biosciences, NJ),mixed, and placed on ice. Plasma was used to determine drug exposure.Liver and quadriceps were removed, immediately freeze-clamped, wrappedin foil and stored in liquid nitrogen.

Tissues were pulverized under liquid N₂ to ensure uniformity insampling. Malonyl-CoA was extracted from the tissue (150-200 mg) with 5volumes 10% tricarboxylic acid in Lysing Matrix A (MP Biomedicals, PN6910) in a FastPrep FP120 (Thermo Scientific, speed=5.5; for 45seconds). The supernatant containing malonyl-CoA was removed from thecell debris after centrifugation at 15000×g for 30 minutes (EppendorfCentrifuge 5402). Samples were stably frozen at −80 C until analysis iscompleted.

Analysis of malonyl CoA levels in liver and muscle tissue can beevaluated using the following methodology.

The method utilizes the following materials: Malonyl-CoA tetralithiumsalt and malonyl-¹³C₃-CoA trilithium salt which were purchased fromIsotec (Miamisburg, Ohio, USA), sodium perchlorate (Sigma, cat no.410241), trichloroacetic acid (ACROS, cat no. 42145), phosphoric acid(J. T. Baker, cat no. 0260-01), ammonium formate (Fluka, cat no. 17843),methanol (HPLC grade, J. T. Baker, cat no. 9093-33), and water (HPLCgrade, J. T. Baker, 4218-03) were used to make the necessary mobilephases. Strata-X on-line solid phase extraction columns, 25 μm, 20mm×2.0 mm I.D (cat no. 00M-S033-B0-CB) were obtained from Phenomenex(Torrance, Calif., USA). SunFire C18 reversed-phase columns, 3.5 μm, 100mm×3.0 mm I.D. (cat no. 186002543) were purchased from WatersCorporation (Milford, Mass., USA).

This method may be performed utilizing the following equipment.Two-dimensional chromatography using an Agilent 1100 binary pump, anAgilent 1100 quaternary pump and two Valco Cheminert 6-port two positionvalves. Samples were introduced via a LEAP HTC PAL auto sampler withPeltier cooled stack maintained at 10° C. and a 20 μL sampling loop. Theneedle wash solutions for the autosampler are 10% trichloroacetic acidin water (w/v) for Wash 1 and 90:10 methanol:water for Wash 2. Theanalytical column (Sunfire) was maintained at 35° C. using a MicroTechScientific Micro-LC Column Oven. The eluent was analyzed on an ABI SciexAPI3000 triple quadrupole mass spectrometer with Turbo Ion Spray.

Two-dimensional chromatography was performed in parallel using distinctgradient elution conditions for on-line solid phase extraction andreversed-phase chromatography. The general design of the method was suchthat the first dimension was utilized for sample clean-up and capture ofthe analyte of interest followed by a brief coupling of both dimensionsfor elution from the first dimension onto the second dimension. Thedimensions were subsequently uncoupled allowing for gradient elution ofthe analyte from the second dimension for quantification whilesimultaneously preparing the first dimension for the next sample in thesequence. When both dimensions were briefly coupled together, the flowof the mobile phase in the first dimension was reversed for analyteelution on to the second dimension, allowing for optimal peak width,peak shape, and elution time.

The first dimension of the HPLC system utilized the Phenomenex strata-Xon-line solid phase extraction column and the mobile phase consisted of100 mM sodium perchlorate/0.1% (v/v) phosphoric acid for solvent A andmethanol for solvent B.

The second dimension of the HPLC system utilized the Waters SunFire C18reversed-phase column and the mobile phase consisted of 100 mM ammoniumformate for solvent A and methanol for solvent B. The initial conditionof the gradient was maintained for 2 minutes and during this time theanalyte was transferred to the analytical column. It was important thatthe initial condition was at a sufficient strength to elute the analytefrom the on-line SPE column while retaining it on the analytical.Afterwards, the gradient rose linearly to 74.5% A in 4.5 minutes beforea wash and re-equilibration step.

Mass spectrometry when coupled with HPLC can be a highly selective andsensitive method for quantitatively measuring analytes in complexmatrices but is still subject to interferences and suppression. Bycoupling a two dimensional HPLC to the mass spectrometer, theseinterferences were significantly reduced. Additionally, by utilizing theMultiple Reaction Monitoring (MRM) feature of the triple quadrupole massspectrometer, the signal-to-noise ratio was significantly improved.

For this assay, the mass spectrometer was operated in positive ion modewith a TurbolonSpray voltage of 2250V. The nebulizing gas was heated to450° C. The Declustering Potential (DP), Focusing Potential (FP), andCollision Energy (CE) were set to 60, 340, and 42 V, respectively.Quadrupole 1 (Q1) resolution was set to unit resolution with Quadrupole3 (Q3) set to low. The CAD gas was set to 8. The MRM transitionsmonitored were for malonyl CoA: 854.1-+347.0 m/z (L. Gao et al. (2007)J. Chromatogr. B 853, 303-313); and for malonyl-¹³C₃-CoA: 857.1→350.0m/z with dwell times of 200 ms. The eluent was diverted to the massspectrometer near the expected elution time for the analyte, otherwiseit was diverted to waste to help preserve the source and improverobustness of the instrumentation. The resulting chromatograms wereintegrated using Analyst software (Applied Biosystems). Tissueconcentrations for malonyl CoA were calculated from a standard curveprepared in a 10% solution of trichloroacetic acid in water.

Samples comprising the standard curve for the quantification ofmalonyl-CoA in tissue extracts were prepared in 10% (w/v)trichloroacetic acid (TCA) and ranged from 0.01 to 1 pmol/μL.Malonyl-¹³C₃-CoA (final concentration of 0.4 pmol/μL) was added to eachstandard curve component and sample as an internal standard.

Six intra-assay quality controls were prepared; three from a pooledextract prepared from fasted animals and three from a pool made from fedanimals. These were run as independent samples spiked with 0, 0.1 or 0.3pmol/μL ¹²C-malonyl-CoA as well as malonyl-¹³C₃-CoA (0.4 pmol/μL). Eachintra-assay quality control contained 85% of aqueous tissue extract withthe remaining portion contributed by internal standard (0.4 pmol/μL) and¹²C-malonyl-CoA. Inter assay controls were included in each run; theyconsist of one fasted and one fed pooled sample of quadriceps and/or onefasted and one fed pooled sample of liver. All such controls are spikedwith malonyl-¹³C₃-CoA (0.4 pmol/μL).

1. A compound of Formula (I)

wherein R¹ is (C₁-C₆)alkyl, (C₃-C₇)cycloalkyl, tetrahydrofuranyl oroxetanyl; wherein said (C₁-C₆)alkyl is optionally substituted with(C₁-C₃)alkoxy phenyl, or tetrahydrofuranyl; R² is hydrogen, halo,(C₁-C₃)alkyl, or cyano; R³ are each independently hydrogen or(C₁-C₃)alkyl; R⁴ is a (C₆-C₁₀)aryl, 5 to 12 membered heteroaryl or 8 to12 membered fused heterocyclicaryl; wherein said (C₆-C₁₀)aryl, 5 to 12membered heteroaryl or 8 to 12 membered fused heterocyclicaryl are eachoptionally substituted with one to three substituents independentlyselected from (C₁-C₃)alkyl, (C₁-C₃)alkoxy, halo, amino,(C₁-C₃)alkylamino, di(C₁-C₃)alkylamino, hydroxy, cyano, amido, phenyl, 5to 6 membered heteroaryl or 5 to 6 membered heterocyclyl; or apharmaceutically acceptable salt thereof.
 2. The compound of claim 1wherein R¹ is (C₁-C₆)alkyl, (C₃-C₇)cycloalkyl, or tetrahydrofuranyl; andR² is hydrogen or methyl; or a pharmaceutically acceptable salt thereof.3. The compound of claim 2 wherein R¹ is ethyl, isopropyl or t-butyl;each R³ is hydrogen; and R⁴ is phenyl, pyrazolyl, imidazolyl, triazolyl,pyridinyl, pyrimidinyl, indolyl, benzopyrazinyl, benzoimidazolyl,benzoimidazolonyl, pyrrolopyridinyl, pyrrolopyrimidinyl,pyrazolopyridinyl, pyrazolopyrimidinyl, indazolyl, indolinonyl,naphthyridinyl, quinolinyl, quinolinonyl, dihydroquinolinonyl,oxo-dihydroquinolinonyl, isoquinolinyl, isoquinolinonyl,dihydroisoquinonyl or oxo-dihydroisoquinonyl, each optionallysubstituted with one to three substituents independently selected fromfluoro, chloro, methyl, methoxy, amino, methylamino, dimethylamino,amido, cyano, phenyl, imidazolyl, pyrazolyl, triazolyl, pyridinyl ormorpholinyl; or a pharmaceutically acceptable salt thereof.
 4. Thecompound of claim 3 wherein R¹ is isopropyl or t-butyl; R² is hydrogen;or a pharmaceutically acceptable salt thereof.
 5. The compound of claim4 wherein R⁴ is indazolyl, benzoimidazolyl,1-oxo-1,2-dihydroisoquinolinyl, 1H-pyrrolo[3,2-b]pyridinyl,2-oxo-2,3-dihydro-1H-benzo[d]imidazolyl, 1H-pyrazolylphenyl,1H-pyrazolylpyridinyl, or 1H-imidazolylphenyl; each optionallysubstituted with one to two methyl, chloro or fluoro; or apharmaceutically acceptable salt thereof.
 6. A compound selected from1-isopropyl-1′-(1-oxo-1,2-dihydroisoquinoline-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(1H-pyrrolo[3,2-b]pyridine-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(2-methyl-3H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(1H-pyrrolo[3,2-b]pyridine-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(1H-indazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(7-fluoro-1H-indazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(1-methyl-2-oxo-2,3-dihydro-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(7-chloro-2-methyl-3H-benzo[d]imidazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(1H-indazole-6-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(3-(1H-pyrazol-4-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(2-methyl-1H-benzo[d]imidazole-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(1H-indazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(1H-indazole-5-carbonyl)-1-isopropyl-3-methyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(2-(1H-pyrazol-3-yl)pyridine-4-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(3-(1H-pyrazol-3-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-isopropyl-1′-(1H-pyrrolo[3,2-b]pyridine-2-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(4-(1H-imidazol-2-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1-(tert-butyl)-1′-(3-(1H-imidazol-2-yl)benzoyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;or1-(tert-butyl)-1′-(1H-indazole-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;or a pharmaceutically acceptable salt thereof.
 7. The compound of claim6 selected from1-isopropyl-1′-(2-methyl-1H-benzo[d]imidazole-6-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(1H-indazole-6-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;1′-(1H-indazole-5-carbonyl)-1-isopropyl-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-one;or a pharmaceutically acceptable salt thereof.
 8. A pharmaceuticalcomposition comprising a therapeutically effective amount of a compoundof claim 1; or a pharmaceutically acceptable salt thereof and apharmaceutically acceptable excipient, diluent, or carrier.
 9. Thecomposition of claim 8 further comprising at least one additionalanti-diabetic agent.
 10. The composition of claim 9 wherein saidanti-diabetic agent is selected from the group consisting of metformin,acetohexamide, chlorpropamide, diabinese, glibenclamide, glipizide,glyburide, glimepiride, gliclazide, glipentide, gliquidone, glisolamide,tolazamide, tolbutamide, tendamistat, trestatin, acarbose, adiposine,camiglibose, emiglitate, miglitol, voglibose, pradimicin-Q, salbostatin,balaglitazone, ciglitazone, darglitazone, englitazone, isaglitazone,pioglitazone, rosiglitazone, troglitazone, exendin-3, exendin-4,trodusquemine, reservatrol, hyrtiosal extract, sitagliptin,vildagliptin, alogliptin and saxagliptin.
 11. A method for treating ordelaying the progression or onset of Type 2 diabetes in humanscomprising the step of administering to the human in need of suchtreatment a therapeutically effective amount of a compound of claim 1 ora pharmaceutically acceptable salt thereof.
 12. A method for treatingnonalcoholic fatty liver disease (NAFLD) or hepatic insulin resistancein humans comprising the step of administering to an animal the human inneed of such treatment a therapeutically effective amount of a compoundof claim 1 or a pharmaceutically acceptable salt thereof.
 13. A methodfor treating or delaying the progression or onset of Type 2 diabetes inhumans comprising the step of administering to the human in need of suchtreatment a pharmaceutical composition of claim
 8. 14. The method ofclaim 13 wherein the progression or onset of Type 2 diabetes anddiabetes-related disorders is delayed.
 15. A method for treatingnonalcoholic fatty liver disease (NAFLD) or hepatic insulin resistancein humans comprising the step of administering to the human in need ofsuch treatment a pharmaceutical composition of claim
 8. 16. A compoundthat is1-isopropyl-1′-(2-methyl-1H-benzo[d]imidazole-5-carbonyl)-4,6-dihydrospiro[indazole-5,4′-piperidin]-7(1H)-oneor a pharmaceutically acceptable salt thereof.
 17. A compound that is


18. A compound that is

or a pharmaceutically acceptable salt thereof.
 19. A compound that is